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  Subjects -> ENVIRONMENTAL STUDIES (Total: 769 journals)
    - ENVIRONMENTAL STUDIES (706 journals)
    - POLLUTION (21 journals)
    - TOXICOLOGY AND ENVIRONMENTAL SAFETY (34 journals)
    - WASTE MANAGEMENT (8 journals)

ENVIRONMENTAL STUDIES (706 journals)            First | 1 2 3 4 5 6 7 8     

International Gambling Studies     Hybrid Journal   (Followers: 5)
International Innovation - climate     Open Access  
International innovation. Environment     Open Access  
International Journal of Acarology     Hybrid Journal   (Followers: 1)
International Journal of Advancement in Earth and Enviromental Sciences     Open Access   (Followers: 2)
International Journal of African Renaissance Studies - Multi-, Inter- and Transdisciplinarity     Hybrid Journal   (Followers: 2)
International Journal of Agricultural and Environmental Information Systems     Full-text available via subscription   (Followers: 1)
International Journal of Alternative Propulsion     Hybrid Journal   (Followers: 1)
International Journal of Applied Psychoanalytic Studies     Hybrid Journal   (Followers: 2)
International Journal of Chinese Culture and Management     Hybrid Journal   (Followers: 1)
International Journal of Corrosion     Open Access   (Followers: 10)
International Journal of Critical Infrastructures     Hybrid Journal   (Followers: 3)
International Journal of Disaster Risk Reduction     Hybrid Journal   (Followers: 6)
International Journal of Disaster Risk Science     Open Access   (Followers: 8)
International Journal of Ecological Economics and Statistics     Full-text available via subscription   (Followers: 1)
International Journal of Ecology     Open Access   (Followers: 8)
International Journal of Ecology & Development     Full-text available via subscription   (Followers: 1)
International Journal of Energy and Environmental Engineering     Open Access   (Followers: 2)
International Journal of Environment     Open Access   (Followers: 3)
International Journal of Environment and Health     Hybrid Journal   (Followers: 7)
International Journal of Environment and Pollution     Hybrid Journal   (Followers: 6)
International Journal of Environment and Sustainable Development     Hybrid Journal   (Followers: 14)
International Journal of Environment and Waste Management     Hybrid Journal   (Followers: 6)
International Journal of Environment, Workplace and Employment     Hybrid Journal   (Followers: 3)
International Journal of Environmental Engineering     Hybrid Journal   (Followers: 5)
International Journal of Environmental Health Engineering     Open Access  
International Journal of Environmental Health Research     Hybrid Journal   (Followers: 3)
International Journal of Environmental Policy and Decision Making     Hybrid Journal   (Followers: 10)
International Journal of Environmental Protection     Open Access   (Followers: 12)
International Journal of Environmental Research and Public Health     Open Access   (Followers: 14)
International Journal of Environmental Science and Technology     Hybrid Journal   (Followers: 5)
International Journal of Environmental Studies     Hybrid Journal   (Followers: 11)
International Journal of Exergy     Hybrid Journal   (Followers: 4)
International Journal of Forest, Soil and Erosion     Open Access   (Followers: 3)
International Journal of Global Environmental Issues     Hybrid Journal   (Followers: 4)
International Journal of Global Warming     Hybrid Journal   (Followers: 5)
International Journal of Greenhouse Gas Control     Partially Free   (Followers: 6)
International Journal of Health Planning and Management     Hybrid Journal   (Followers: 6)
International Journal of Hygiene and Environmental Health     Hybrid Journal   (Followers: 6)
International Journal of Logistics Research and Applications : A Leading Journal of Supply Chain Management     Hybrid Journal   (Followers: 9)
International Journal of Philosophical Studies     Hybrid Journal   (Followers: 2)
International Journal of Phytoremediation     Hybrid Journal   (Followers: 1)
International Journal of Process Systems Engineering     Hybrid Journal   (Followers: 1)
International Journal of Recycling of Organic Waste in Agriculture     Open Access   (Followers: 1)
International Journal of Regulation and Governance     Hybrid Journal   (Followers: 2)
International Journal of Reliability and Safety     Hybrid Journal   (Followers: 5)
International Journal of Renewable Energy Development     Open Access   (Followers: 6)
International Journal of Social Sciences and Management     Open Access  
International Journal of Soil, Sediment and Water     Open Access   (Followers: 8)
International Journal of Stress Management     Full-text available via subscription   (Followers: 8)
International Journal of Sustainable Construction Engineering and Technology     Open Access   (Followers: 7)
International Journal of Sustainable Engineering     Hybrid Journal   (Followers: 7)
International Journal of Sustainable Materials and Structural Systems     Hybrid Journal   (Followers: 5)
International Journal of Sustainable Society     Hybrid Journal   (Followers: 7)
International Journal of Testing     Hybrid Journal   (Followers: 1)
International Journal of the Commons     Open Access   (Followers: 3)
International Journal of Toxicology     Hybrid Journal   (Followers: 6)
International Journal of Water Resources and Environmental Engineering     Open Access   (Followers: 1)
International Review of Environmental and Resource Economics     Full-text available via subscription  
International Studies in the Philosophy of Science     Hybrid Journal   (Followers: 9)
Interventions : International Journal of Postcolonial Studies     Hybrid Journal   (Followers: 5)
IOP Conference Series: Earth and Environmental Science     Open Access   (Followers: 7)
Iranian Journal of Environmental Health Science & Engineering     Open Access   (Followers: 1)
Iranian Studies     Hybrid Journal   (Followers: 7)
Irish Educational Studies     Hybrid Journal   (Followers: 1)
Irish Journal of Earth Sciences     Full-text available via subscription  
Irish Political Studies     Hybrid Journal   (Followers: 7)
ISLE: Interdisciplinary Studies in Literature and Environment     Hybrid Journal   (Followers: 2)
Isotopes in Environmental and Health Studies     Hybrid Journal   (Followers: 2)
Israel Studies     Full-text available via subscription   (Followers: 5)
ISRN Ecology     Open Access  
ISRN Environmental Chemistry     Open Access  
Jahangirnagar University Environmental Bulletin     Open Access  
Journal of Bioremediation & Biodegradation     Open Access   (Followers: 2)
Journal of Earth Science & Climatic Change     Open Access   (Followers: 8)
Journal of Petroleum & Environmental Biotechnology     Open Access   (Followers: 2)
Journal of Advances in Environmental Health Research     Open Access  
Journal of Agricultural and Environmental Ethics     Hybrid Journal   (Followers: 7)
Journal of Agricultural Biotechnology and Sustainable Development     Open Access  
Journal of Agriculture and Environment     Open Access  
Journal of Agriculture and Environment for International Development     Open Access   (Followers: 6)
Journal of Agrobiology     Open Access   (Followers: 2)
Journal of Applied Ecology     Hybrid Journal   (Followers: 196)
Journal of Applied Meteorology and Climatology     Full-text available via subscription   (Followers: 7)
Journal of Applied Psychoanalytic Studies     Hybrid Journal   (Followers: 1)
Journal of Applied Sciences and Environmental Management     Open Access   (Followers: 1)
Journal of Applied Sciences in Environmental Sanitation     Open Access   (Followers: 6)
Journal of Applied Toxicology     Hybrid Journal   (Followers: 9)
Journal of Applied Volcanology     Open Access   (Followers: 6)
Journal of Arid Environments     Hybrid Journal   (Followers: 8)
Journal of Asian Studies     Full-text available via subscription   (Followers: 23)
Journal of Biochemical and Molecular Toxicology     Hybrid Journal   (Followers: 5)
Journal of Black Studies     Hybrid Journal   (Followers: 2)
Journal of Chemical Ecology     Hybrid Journal   (Followers: 2)
Journal of Chemical Health and Safety     Hybrid Journal   (Followers: 2)
Journal of Climate     Full-text available via subscription   (Followers: 24)
Journal of Coastal Research     Full-text available via subscription   (Followers: 11)
Journal of Computational Environmental Sciences     Open Access  
Journal of Contaminant Hydrology     Hybrid Journal   (Followers: 9)
Journal of Contemporary European Studies     Hybrid Journal   (Followers: 4)

  First | 1 2 3 4 5 6 7 8     

Journal Cover Journal of Applied Toxicology
   Journal TOC RSS feeds Export to Zotero [11 followers]  Follow    
   Hybrid Journal Hybrid journal (It can contain Open Access articles)
     ISSN (Print) 0260-437X - ISSN (Online) 1099-1263
     Published by John Wiley and Sons Homepage  [1604 journals]   [SJR: 0.689]   [H-I: 47]
  • Mini review on blood–brain barrier penetration of pyridinium
           aldoximes
    • Authors: H. Kalász; S. M. Nurulain, G. Veress, S. Antus, F. Darvas, E. Adeghate, A. Adem, F. Hashemi, K. Tekes
      Pages: n/a - n/a
      Abstract: This paper reviews the blood–brain barrier (BBB) penetration of newly developed pyridinium aldoximes. Pyridinium aldoximes are highly charged hydrophilic compounds used in the treatment of subjects exposed to organophosphonates because they are effective as acetylcholinesterase reactivators. Pyridinium aldoximes have antidotal effects against poisoning with cholinesterase inhibitors, a frequent problem affecting people working with organophosphate‐based insecticides and pesticides. Toxic organophosphonate products such as sarin and tabun can be used by terrorists as chemical warfare agents. This poses a severe challenge to all innocent and peace‐loving people worldwide. This review gives a brief summary of BBB transporters and description of the current in vitro and in vivo methods for the characterization of BBB penetration of established and novel pyridinium aldoximes. The authors provide a putative mechanism of penetration, outline some future ways of formulation and discuss the possible advantages and disadvantages of increasing BBB penetration. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-10-07T05:21:33.385324-05:
      DOI: 10.1002/jat.3048
       
  • Neurotoxic effects of ochratoxin A on the subventricular zone of adult
           mouse brain
    • Authors: Sara Paradells; Brenda Rocamonde, Cristina Llinares, Vicente Herranz‐Pérez, Misericordia Jimenez, Jose Manuel Garcia‐Verdugo, Ivan Zipancic, Jose Miguel Soria, Ma. Angeles Garcia‐Esparza
      Abstract: Ochratoxin A (OTA), a mycotoxin that was discovered as a secondary metabolite of the fungal species Aspergillus and Penicillium, is a common contaminant in food and animal feed. This mycotoxin has been described as teratogenic, carcinogenic, genotoxic, immunotoxic and has been proven a potent neurotoxin. Other authors have previously reported the effects of OTA in different structures of the central nervous system as well as in some neurogenic regions. However, the impact of OTA exposure in the subventricular zone (SVZ) has not been assessed yet. To elucidate whether OTA affects neural precursors of the mouse SVZ we investigated, in vitro and in vivo, the effects of OTA exposure on the SVZ and on the neural precursors obtained from this neurogenic niche. In this work, we prove the cumulative effect of OTA exposure on proliferation, differentiation and depletion of neural stem cells cultured from the SVZ. In addition, we corroborated these results in vivo by immunohistochemistry and electron microscopy. As a result, we found a significant alteration in the proliferation process, which was evidenced by a decrease in the number of 5‐bromo‐2‐deoxyuridine‐positive cells and glial cells, as well as, a significant decrease in the number of neuroblasts in the SVZ. To summarize, in this study we demonstrate how OTA could be a threat to the developing and the adult SVZ through its impact in cell viability, proliferation and differentiation in a dose‐dependent manner. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-25T07:57:47.345347-05:
      DOI: 10.1002/jat.3061
       
  • Oral cadmium exposure during rat pregnancy: assessment of transplacental
           micronutrient transport and steroidogenesis at term
    • Authors: Anja Mikolić; Martina Piasek, Antonija Sulimanec Grgec, Veda M. Varnai, Sandra Stasenko, Saša Kralik Oguić
      Abstract: Diet is the main source of cadmium (Cd) exposure. Gastrointestinal absorption increases during pregnancy. Cadmium accumulated in the placenta may interfere with nutrient transport to the foetus. Data on the potential of Cd to act as a steroid disruptor of pregnancy are limited. We evaluated the effects of oral Cd exposure during pregnancy on placental function in micronutrient transfer to the foetus and steroidogenesis in Wistar rats (regular 4‐day cyclers) that mated with unexposed males. Pregnant rats were randomly assigned to a Cd group exposed orally to 50 mg Cd l–1 (CdCl2xH2O dissolved in demineralized water), ≈7.5 mg Cd kg–1 a day, during 20 days of gestation and control (supplied with demineralized water). Non‐pregnant rats were treated under the same experimental conditions. On day 20, all of the rats were killed and samples were taken for element analyses (by electrothermal atomic absorption spectrometry). Progesterone and testosterone were measured in serum and placenta‐derived samples (by immunoenzymometric assay and/or enzyme‐linked immunosorbent assay). In the exposed rats, Cd increased in blood and organs, more in pregnant rats, and in placenta and foetus whereas zinc increased in liver. Iron decreased in maternal organs and in foetus, whereas zinc decreased in maternal kidney and placenta. Liver copper was lower and kidney copper higher in all pregnant vs. non‐pregnant rats. Steroids in serum and placenta did not change. In conclusion, oral Cd exposure during rat pregnancy does not affect progesterone and testosterone at term. Transplacental iron and zinc handover are disrupted, which may put at risk the maintenance of foetal nutrition and viability. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-25T07:52:07.664676-05:
      DOI: 10.1002/jat.3055
       
  • Evaluation and refinement of a field‐portable drinking water
           toxicity sensor utilizing electric cell–substrate impedance sensing
           and a fluidic biochip
    • Authors: Mark W. Widder; Linda M. Brennan, Elizabeth A. Hanft, Mary E. Schrock, Ryan R. James, William H. Schalie
      Abstract: The US Army's need for a reliable and field‐portable drinking water toxicity sensor was the catalyst for the development and evaluation of an electric cell–substrate impedance sensing (ECIS) device. Water testing technologies currently available to soldiers in the field are analyte‐specific and have limited capabilities to detect broad‐based water toxicity. The ECIS sensor described here uses rainbow trout gill epithelial cells seeded on fluidic biochips to measure changes in impedance for the detection of possible chemical contamination of drinking water supplies. Chemicals selected for testing were chosen as representatives of a broad spectrum of toxic industrial compounds. Results of a US Environmental Protection Agency (USEPA)‐sponsored evaluation of the field portable device were similar to previously published US Army testing results of a laboratory‐based version of the same technology. Twelve of the 18 chemicals tested following USEPA Technology Testing and Evaluation Program procedures were detected by the ECIS sensor within 1 h at USEPA‐derived human lethal concentrations. To simplify field‐testing methods further, elimination of a procedural step that acclimated cells to serum‐free media streamlined the test process with only a slight loss of chemical sensitivity. For field use, the ECIS sensor will be used in conjunction with an enzyme‐based sensor that is responsive to carbamate and organophosphorus pesticides. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-18T03:31:57.64557-05:0
      DOI: 10.1002/jat.3017
       
  • Characteristic molecular and proteomic signatures of drug‐induced
           liver injury in a rat model
    • Authors: Jung Woo Eun; Hyun Jin Bae, Qingyu Shen, Se Jin Park, Hyung Seok Kim, Woo Chan Shin, Hee Doo Yang, Chan Young Jin, Jueng Soo You, Hyun Joo Kang, Hoguen Kim, Young Min Ahn, Won Sang Park, Jung Young Lee, Suk Woo Nam
      Abstract: Drug‐induced liver injury (DILI) is a major safety concern during drug development and remains one of the main reasons for withdrawal of drugs from the market. Although it is crucial to develop methods that will detect potential hepatotoxicity of drug candidates as early and as quickly as possible, there is still a lack of sensitive and specific biomarkers for DILI that consequently leads to a scarcity of reliable hepatotoxic data. Hence, in this study, we assessed characteristic molecular signatures in rat liver treated with drugs (pyrazinamide, ranitidine, enalapril, carbamazepine and chlorpromazine) that are known to cause DILI in humans. Unsupervised hierarchical clustering analysis of transcriptome changes induced by DILI‐causing drugs resulted in three different subclusters on dendrogram, i.e., hepatocellular, cholestatic and mixed type of DILI at early time points (2 days), and multiclassification analysis suggested 31 genes as discernible markers for each DILI pattern. Further analysis for characteristic molecular signature of each DILI pattern provided a molecular basis for different modes of DILI action. A proteomics study of the same rat livers was used to confirm the results, and the two sets of data showed 60 matching classifiers. In conclusion, the data of different DILI‐causing drug treatments from genomic analysis in a rat model suggest that DILI‐specific molecular signatures can discriminate different patterns of DILI at an early exposure time point, and that they provide useful information for mechanistic studies that may lead to a better understanding of the molecular basis of DILI. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-18T03:11:37.586599-05:
      DOI: 10.1002/jat.3062
       
  • Chronic trimethyltin chloride exposure and the development of kidney
           stones in rats
    • Authors: Xuefeng Ren; Xin Wu, Gang Sui, Zhihong Gong, Emmanuel Yawson, Banghua Wu, Guanchao Lai, Xiaolin Ruan, Hongbin Gao, Feng Zhou, Bing Su, James R. Olson, Xiaojiang Tang
      Abstract: We recently reported that occupational exposure to trimethyltin (TMT) is a risk factor for developing kidney stones. To further examine the association between TMT exposure and the formation of kidney stones, we conducted a 180‐day animal study and exposed the randomly grouped Sprague–Dawley (SD) rats to TMT in the drinking water at doses of 0, 8.2, 32.8 and 131.3 µg kg–1 day–1. Transient behavioral changes were observed in the high‐dose group during the first 2 weeks of exposure. TMT exposure led to a significant dose‐dependent inhibition of renal H+/K+‐ATPase and an increase in urinary pH. In comparison to no kidney stones being identified in the control and the lowest dose group, 1 rat in the 32.8 µg kg–1 day–1 dose group and 3 out of 9 rats in the 131.3 µg kg–1 day–1 dose group were found to have stones in the kidney/urinary tract. Pathological analysis showed that more wide spread calcium disposition was observed in kidneys of rats with TMT exposure compared with the rats in the control group. However, X‐ray diffraction (XRD) analysis found that the kidney stones were mainly composed of struvite with the formula: NH4MgPO4 6H2O, while calcium‐containing components were also detected. Together, this study further demonstrates through animal studies that chronic exposure to a relatively low level of TMT induces nephrotoxicity and increases the risk for developing kidney stones. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-16T05:47:12.006379-05:
      DOI: 10.1002/jat.3054
       
  • Non‐clinical safety evaluation of single and repeated intramuscular
           administrations of MAGE‐A3 Cancer Immunotherapeutic in rabbits and
           cynomolgus monkeys
    • Authors: Eric Destexhe; Emilie Grosdidier, Nathalie Baudson, Roy Forster, Catherine Gerard, Nathalie Garçon, Lawrence Segal
      Abstract: The MAGE‐A3 recombinant protein combined with AS15 immunostimulant (MAGE‐A3 Cancer Immunotherapeutic) is under development by GlaxoSmithKline for the treatment of lung cancer and melanoma. We performed non‐clinical safety studies evaluating potential local and systemic toxic effects induced by MAGE‐A3 Cancer Immunotherapeutic in rabbits (study 1) and cynomolgus monkeys (study 2). Animals were allocated to two groups to receive a single (rabbits) or 25 repeated (every 2 weeks) injections (monkeys) of MAGE‐A3 Cancer Immunotherapeutic (treatment groups) or saline (control groups). All rabbits were sacrificed 3 days post‐injection and monkeys 3 days following last injection (3/5 per gender per group) or after a 3‐month treatment‐free period (2/5 per gender per group). Local and systemic reactions and MAGE‐A3‐specific immune responses (monkeys) were assessed. Macroscopic and microscopic (for rabbits, injection site only) post‐mortem examinations were performed on all animals. No systemic toxicity or unscheduled mortalities were recorded. Single (rabbits) and repeated (monkeys; up to four times at the same site) injections were well tolerated. Following five to seven repeated injections, limb circumferences increased up to 26% (5 h post‐injection), but returned to normal after 1–8 days. Three days after the last injection, enlargements of iliac, popliteal, axillary and inguinal lymph nodes, and increased incidence or severity of mononuclear inflammatory cell infiltrates was observed in injected muscles of treated monkeys. No treatment‐related macroscopic findings were recorded after the treatment‐free period. MAGE‐A3‐specific antibody and T‐cell responses were raised in all treated monkeys, confirming test item exposure. Single or repeated intramuscular injections of MAGE‐A3 Cancer Immunotherapeutic were well tolerated in rabbits and monkeys. Copyright © 2014 GlaxoSmithKline Vaccines. Journal of Applied Toxicology published by John Wiley & Sons, Ltd.
      PubDate: 2014-09-12T22:50:49.939421-05:
      DOI: 10.1002/jat.3025
       
  • Graphene supports in vitro proliferation and osteogenic differentiation of
           goat adult mesenchymal stem cells: potential for bone tissue engineering
    • Authors: Hoda Elkhenany; Lisa Amelse, Andersen Lafont, Shawn Bourdo, Marc Caldwell, Nancy Neilsen, Enkeleda Dervishi, Oshin Derek, Alexandru S. Biris, David Anderson, Madhu Dhar
      Abstract: Current treatments for bone loss injuries involve autologous and allogenic bone grafts, metal alloys and ceramics. Although these therapies have proved useful, they suffer from inherent challenges, and hence, an adequate bone replacement therapy has not yet been found. We hypothesize that graphene may be a useful nanoscaffold for mesenchymal stem cells and will promote proliferation and differentiation into bone progenitor cells. In this study, we evaluate graphene, a biocompatible inert nanomaterial, for its effect on in vitro growth and differentiation of goat adult mesenchymal stem cells. Cell proliferation and differentiation are compared between polystyrene‐coated tissue culture plates and graphene‐coated plates. Graphitic materials are cytocompatible and support cell adhesion and proliferation. Importantly, cells seeded on to oxidized graphene films undergo osteogenic differentiation in fetal bovine serum‐containing medium without the addition of any glucocorticoid or specific growth factors. These findings support graphene's potential to act as an osteoinducer and a vehicle to deliver mesenchymal stem cells, and suggest that the combination of graphene and goat mesenchymal stem cells provides a promising construct for bone tissue engineering. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-12T22:12:52.239351-05:
      DOI: 10.1002/jat.3024
       
  • Exposure to MnCl2 · 4H2O during development induces
           activation of microglial and perivascular macrophage populations in the
           hippocampal dentate gyrus of rats
    • Authors: Hajime Abe; Takumi Ohishi, Fumiyuki Nakane, Ayako Shiraki, Takeshi Tanaka, Toshinori Yoshida, Makoto Shibutani
      Abstract: Developmental exposure to Mn caused Mn accumulation in the brain tissue and transient disruption of granule cell neurogenesis, targeting the late stage differentiation of progenitor cells in the subgranular zone of the hippocampal dentate gyrus of rats. Because neurogenesis is influenced by proinflammatory responses, this study was performed to determine whether Mn exposure causes microglial activation in the dentate hilus, a region anatomically close to the subgranular zone of the dentate gyrus. Pregnant rats were treated with dietary MnCl2 · 4H2O at 32, 160 or 800 ppm from gestational day 10 to day 21 after delivery. An immunohistochemical analysis revealed increases in Iba1+ microglia in the hilus on postnatal day 21 following exposure to MnCl2 · 4H2O in a dose‐unrelated manner at 32 and at 800 ppm and an increase in CD163+ macrophage at 800 ppm in the hilus. Real‐time reverse transcription–polymerase chain reaction analysis revealed increases in the mRNA levels of Il1α, Il6, Nos2 and Tnf after 800 ppm MnCl2 · 4H2O. These results suggest that activation of microglia and perivascular macrophages occurs in the hilus after developmental exposure to MnCl2 · 4H2O at 800 ppm, and probably involves the disruption of neurogenesis through the accumulation of Mn in the brain tissue. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-12T22:06:33.032918-05:
      DOI: 10.1002/jat.3059
       
  • Cylindrospermopsin induces oxidative stress and genotoxic effects in the
           fish CLC cell line
    • Authors: Anna Sieroslawska; Anna Rymuszka
      Abstract: Cylindrospermopsin (CYN) is a cyanotoxin detected in water reservoirs worldwide. The toxin is a potent protein synthesis inhibitor capable of adversely influencing a wide range of cell functions. While data on the prooxidative potency of CYN are inconsistent, genotoxic effects towards certain mammalian cell types have been described. However, such a potential on fish cells has not yet been investigated. Hence, the aim of the study was to elucidate the prooxidative and genotoxic impact of CYN on a common carp (Cyprinus carpio L.) leucocyte cell line (CLC). The cells were incubated with the cyanotoxin at concentrations of 0.1, 0.5 or 1 µg ml–1. After 24 h, cytotoxic activity of CYN at the highest used concentration was confirmed by decreased cell membrane integrity and inhibited cell proliferation. Additionally, CYN at 0.5 and 1 µg ml–1 increased intracellular ATP levels and decreased the reduced to oxidized glutathione ratio. Furthermore, a significant increase in the reactive oxygen species (ROS) production with concomitant changes in superoxide dismutase activity was observed after a 3.5‐h exposure of the cells to the toxin. Genotoxic activity of CYN, manifested as oxidative DNA damage and elevated number of micronuclei, was also detected in exposed cells. The obtained results indicate that CYN is able to exert a wide range of adverse effects, including oxidative stress and genotoxicity in fish leucocytes. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-12T22:06:25.237034-05:
      DOI: 10.1002/jat.3040
       
  • Development of haemostatic decontaminants for the treatment of wounds
           contaminated with chemical warfare agents. 2: Evaluation of in vitro
           topical decontamination efficacy using undamaged skin
    • Authors: Christopher H. Dalton; Charlotte A. Hall, Helen L. Lydon, J. K. Chipman, John S. Graham, John Jenner, Robert P. Chilcott
      Abstract: The risk of penetrating, traumatic injury occurring in a chemically contaminated environment cannot be discounted. Should a traumatic injury be contaminated with a chemical warfare (CW) agent, it is likely that standard haemostatic treatment options would be complicated by the need to decontaminate the wound milieu. Thus, there is a need to develop haemostatic products that can simultaneously arrest haemorrhage and decontaminate CW agents. The purpose of this study was to evaluate a number of candidate haemostats for efficacy as skin decontaminants against three CW agents (soman, VX and sulphur mustard) using an in vitro diffusion cell containing undamaged pig skin. One haemostatic product (WoundStat™) was shown to be as effective as the standard military decontaminants Fuller's earth and M291 for the decontamination of all three CW agents. The most effective haemostatic agents were powder‐based and use fluid absorption as a mechanism of action to sequester CW agent (akin to the decontaminant Fuller's earth). The envisaged use of haemostatic decontaminants would be to decontaminate from within wounds and from damaged skin. Therefore, WoundStat™ should be subject to further evaluation using an in vitro model of damaged skin. Copyright © 2014 Crown copyright. Journal of Applied Toxicology © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-12T21:56:06.175164-05:
      DOI: 10.1002/jat.3060
       
  • Early chronic lead exposure reduces exploratory activity in young C57BL/6J
           mice
    • Authors: Mayra Gisel Flores‐Montoya; Christina Sobin
      Abstract: Research has suggested that chronic low‐level lead exposure diminishes neurocognitive function in children. Tests that are sensitive to behavioral effects at lowest levels of lead exposure are needed for the development of animal models. In this study we investigated the effects of chronic low‐level lead exposure on exploratory activity (unbaited nose poke task), exploratory ambulation (open field task) and motor coordination (Rotarod task) in pre‐adolescent mice. C57BL/6J pups were exposed to 0 ppm (controls), 30 ppm (low‐dose) or 230 ppm (high‐dose) lead acetate via dams’ drinking water administered from birth to postnatal day 28, to achieve a range of blood lead levels (BLLs) from not detectable to 14.84 µg dl–1). At postnatal day 28, mice completed behavioral testing and were killed (n = 61). BLLs were determined by inductively coupled plasma mass spectrometry. The effects of lead exposure on behavior were tested using generalized linear mixed model analyses with BLL, sex and the interaction as fixed effects, and litter as the random effect. BLL predicted decreased exploratory activity and no threshold of effect was apparent. As BLL increased, nose pokes decreased. The C57BL/6J mouse is a useful model for examining effects of early chronic low‐level lead exposure on behavior. In the C57BL/6J mouse, the unbaited nose poke task is sensitive to the effects of early chronic low‐level lead exposure. This is the first animal study to show behavioral effects in pre‐adolescent lead‐exposed mice with BLL below 5 µg dl–1. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-12T21:41:49.427385-05:
      DOI: 10.1002/jat.3064
       
  • Diagnostic and predictive performance and standardized threshold of
           traditional biomarkers for drug‐induced liver injury in rats
    • Authors: Yutaka Tonomura; Yuki Kato, Hiroyuki Hanafusa, Yuji Morikawa, Keigo Matsuyama, Takeki Uehara, Motonobu Ueno, Mikinori Torii
      Pages: n/a - n/a
      Abstract: Traditional biomarkers such as alanine and aspartate aminotransferase (ALT, AST) and total bilirubin (TBIL) have been widely used for detecting drug‐induced liver injury (DILI). Although the Food and Drug Administration (FDA) proposed standardized thresholds for human as Hy's law, those for animals have not been determined, and predictability of these biomarkers for future onset of hepatic lesions remains unclear. In this study, we investigated these diagnostic and predictive performance of 10 traditional biomarkers for liver injury by receiver‐operating characteristic (ROC) curve, using a free‐access database where 142 hepatotoxic or non‐hepatotoxic compounds were administrated to male rats (n = 5253). Standardization of each biomarker value was achieved by calculating the ratio to control mean value, and the thresholds were determined under the condition of permitting 5% false positive. Of these 10 biomarkers, AST showed the best diagnostic performance. Furthermore, ALT and TBIL also showed high performance under the situation of hepatocellular necrosis and bile duct injury, respectively. Additionally, the availability of the diagnostic thresholds in difference testing facility was confirmed by the application of these thresholds to in‐house prepared dataset. Meanwhile, incorrect diagnosis by the thresholds was also observed. Regarding prediction, all 10 biomarkers showed insufficient performance for future onset of hepatic lesions. In conclusion, the standardized diagnostic thresholds enable consistent evaluation of traditional biomarkers among different facilities, whereas it was suggested that novel biomarker is required for more accurate diagnosis and prediction of DILI. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-04T05:02:30.374456-05:
      DOI: 10.1002/jat.3053
       
  • Perturbation of cytosolic calcium by 2‐aminoethoxydiphenyl borate
           and caffeine affects zebrafish myofibril alignment
    • Authors: Hsin‐Ju Wu; Tsorng‐Harn Fong, Shen‐Liang Chen, Jen‐Cheng Wei, I‐Jong Wang, Chi‐Chung Wen, Chao‐Yuan Chang, Xing‐Guang Chen, Wei‐Yu Chen, Hui‐Min Chen, Juin‐Lin Horng, Yun‐Hsin Wang, Yau‐Hung Chen
      Pages: n/a - n/a
      Abstract: The objective of the current study was to investigate the effects of Ca2+ levels on myofibril alignment during zebrafish embryogenesis. To investigate how altered cytoplasmic Ca2+ levels affect myofibril alignment, we exposed zebrafish embryos to 2‐aminothoxyldiphenyl borate (2‐APB; an inositol 1,4,5‐trisphosphate receptor inhibitor that reduces cytosolic Ca2+ levels) and caffeine (a ryanodine receptor activator that enhances cytosolic Ca2+ levels). The results demonstrated that the most evident changes in zebrafish embryos treated with 2‐APB were shorter body length, curved trunk and malformed somite boundary. In contrast, such malformed phenotypes were evident neither in untreated controls nor in caffeine‐treated embryos. Subtle morphological changes, including changes in muscle fibers, F‐actin and ultrastructures were easily observed by staining with specific monoclonal antibodies (F59 and α‐laminin), fluorescent probes (phalloidin) and by transmission electron microscopy. Our data suggested that: (1) the exposure to 2‐APB and/or caffeine led to myofibril misalignment; (2) 2‐APB‐treated embryos displayed split and short myofibril phenotypes, whereas muscle fibers from caffeine‐treated embryos were twisted and wavy; and (3) zebrafish embryos co‐exposed to 2‐APB and caffeine resulted in normal myofibril alignment. In conclusion, we proposed that cytosolic Ca2+ is important for myogenesis, particularly for myofibril alignment. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-04T04:56:51.010578-05:
      DOI: 10.1002/jat.3057
       
  • Skin absorption of six performance amines used in metalworking fluids
    • Authors: Lauriane N. Roux; James D. Brooks, James L. Yeatts, Ronald E. Baynes
      Pages: n/a - n/a
      Abstract: Every year, 10 million workers are exposed to metalworking fluids (MWFs) that may be toxic. There are four types of MWFs: neat oils and three water‐based MWFs (soluble oil, semisynthetic and synthetic), which are diluted with water and whose composition varies according to the mineral oils ratio. MWFs also contain various additives. To determine the absorption of six amines used as corrosion inhibitors and biocides in MWFs, porcine skin flow‐through diffusion cell experiments were conducted with hydrophilic ethanolamines (mono‐, di‐ and triethanolamine, MEA, DEA and TEA respectively) and a mixture of lipophilic amines (dibutylethanolamine, dicyclohexylamine and diphenylamine). The six amines were dosed in four vehicles (water and three generic water‐based MWF formulations) and analyzed using a scintillation counter or gas chromatography/mass spectrometry. These 24 h studies showed that dermal absorption significantly (P 
      PubDate: 2014-09-04T04:56:30.081537-05:
      DOI: 10.1002/jat.3056
       
  • A representative retinoid X receptor antagonist UVI3003 induced
           teratogenesis in zebrafish embryos
    • Authors: Liang Zheng; Ting Xu, Daoji Li, Junliang Zhou
      Pages: n/a - n/a
      Abstract: Retinoid X receptor (RXR) interfering activity has been detected in different water resources. To study RXR disruptor‐induced toxicological effects on vertebrates, embryos of zebrafish (Danio rerio) were exposed to a representative RXR antagonist UVI3003. Results showed that the teratogenic index (LC50/EC50) of UVI3003 was as high as 5.4. UVI3003 induced multiple malformations of embryos, including deformed fins, reduced brains, small jaws, bent tails and edema in hearts, the degree of which became more severe with increasing exposure concentration. Although no significant difference was observed in the hatching rates between the exposure group and control, the whole body length was significantly reduced by 6.5% and 8.9% when exposed to 200 and 300 µg l−1 of UVI3003, respectively. The heart rate also significantly decreased by 8.8–50.2% during exposure. Further experiments revealed that the pharyngula stage was the most sensitive development phase in terms of embryo response to UVI3003. The results demonstrated severe teratogenicity of RXR antagonist in zebrafish embryos and provided important data for ecotoxicological evaluation of RXR antagonists. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-03T03:07:21.862962-05:
      DOI: 10.1002/jat.3051
       
  • Immunophenotyping does not improve predictivity of the local lymph node
           assay in mice
    • Authors: Volker Strauss; Susanne N. Kolle, Naveed Honarvar, Martina Dammann, Sibylle Groeters, Frank Faulhammer, Robert Landsiedel, Bennard Ravenzwaay
      Pages: n/a - n/a
      Abstract: The local lymph node assay (LLNA) is a regulatory accepted test for the identification of skin sensitizing substances by measuring radioactive thymidine incorporation into the lymph node. However, there is evidence that LLNA is overestimating the sensitization potential of certain substance classes in particular those exerting skin irritation. Some reports describe the additional use of flow cytometry‐based immunophenotyping to better discriminate irritants from sensitizing irritants in LLNA. In the present study, the 22 performance standards plus 8 surfactants were assessed using the radioactive LLNA method. In addition, lymph node cells were immunophenotyped to evaluate the specificity of the lymph node response using cell surface markers such as B220 or CD19, CD3, CD4, CD8, I‐Aκ and CD69 with the aim to allow a better discrimination above all between irritants and sensitizers, but also non‐irritating sensitizers and non‐sensitizers. However, the markers assessed in this study do not sufficiently differentiate between irritants and irritant sensitizers and therefore did not improve the predictive capacity of the LLNA. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-03T03:06:12.16689-05:0
      DOI: 10.1002/jat.3042
       
  • Reversible cholinesterase inhibitors as pre‐treatment for exposure
           to organophosphates: assessment using azinphos‐methyl
    • Authors: Georg A. Petroianu; Syed M. Nurulain, Mohamed Y. Hasan, Kamil Kuča, Dietrich E. Lorke
      Pages: n/a - n/a
      Abstract: Pre‐treatment with reversible acetylcholinesterase (AChE) inhibitors before organophosphorous compound (OPC) exposure can reduce OPC‐induced mortality. However, pyridostigmine, the only substance employed for such prophylaxis, is merely efficacious against a limited number of OPCs. In search of more efficacious and broad‐range alternatives, we have compared in vivo the ability of five reversible AChE inhibitors (pyridostigmine, physostigmine, ranitidine, tacrine and K‐27) to reduce mortality induced by the OPC azinphos‐methyl. Protection was quantified using Cox analysis by determining the relative risk (RR) of death in rats that were administered these AChE inhibitors in equitoxic dosage (25% of LD01) 30 min before azinphos‐methyl exposure. Azinphos‐methyl‐induced mortality was significantly reduced by all five tested compounds as compared with the reference group that was only exposed to azinphos‐methyl without prior pre‐treatment (RR = 1). The most efficacious prophylactic agents were K‐27 (RR = 0.15) and physostigmine (RR = 0.21), being significantly more efficacious than ranitidine (RR = 0.62) and pyridostigmine (RR = 0.37). Pre‐treatment with tacrine (RR = 0.29) was significantly more efficacious than pre‐treatment with ranitidine, but the difference between tacrine and pyridostigmine was not significant. Our results indicate that prophylactic administration of the oxime K‐27 may be a promising alternative in cases of imminent OPC exposure. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-03T03:04:57.769676-05:
      DOI: 10.1002/jat.3052
       
  • Carnosic acid induces autophagic cell death through inhibition of the
           Akt/mTOR pathway in human hepatoma cells
    • Authors: Qilong Gao; Huaimin Liu, Yamin Yao, Liang Geng, Xinfeng Zhang, Lifeng Jiang, Bian Shi, Feng Yang
      Abstract: The therapeutic goal of cancer treatment is now geared towards triggering tumour‐selective cell death with autophagic cell death being required for the chemotherapy of apoptosis‐resistant cancer. In this study, Carnosic acid (CA), a polyphenolic diterpene isolated from Rosemary (Rosemarinus officinalis), significantly induced autophagic cell death in HepG2 cells. Ca treatment caused the formation of autophagic vacuoles produced an increasing ratio of LC3‐II to LC3‐I in a time‐ and dose‐dependent manner but had no effect on the levels of autophagy‐related protein ATG6 and ATG13 expression. Autophagy inhibitors, 3‐methyladenine (3‐MA), chloroquine and bafilomycin A1, or ATG genes silencing in HepG2 cells significantly inhibited CA‐induced autophagic cell death. The CA treatment decreased the levels of phosphorylated Akt and mTOR without any effects on PI3K or PTEN. Most importantly, overexpression of Akt and knockdown of PTEN attenuated autophagy induction in CA‐treated cells. Taken together, our results indicated that CA induced autophagic cell death through inhibition of the Akt/mTOR pathway in human hepatoma cells. These findings suggest that CA has a great potential for the treatment of hepatoma via autophagic induction. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-01T07:01:21.780556-05:
      DOI: 10.1002/jat.3049
       
  • Protective effects of ascorbic acid against the genetic and epigenetic
           alterations induced by 3,5‐dimethylaminophenol in AA8 cells
    • Authors: Ming‐Wei Chao; P𝚤nar Erkekoglu, Chia‐Yi Tseng, Wenjie Ye, Laura J. Trudel, Paul L. Skipper, Steven R. Tannenbaum, Gerald N. Wogan
      Abstract: Exposure to monocyclic aromatic alkylanilines (MAAs), namely 2,6‐dimethylaniline (2,6‐DMA), 3,5‐dimethylaniline (3,5‐DMA) and 3‐ethylaniline (3‐EA), was significantly and independently associated with bladder cancer incidence. 3,5‐DMAP (3,5‐dimethylaminophenol), a metabolite of 3,5‐DMA, was shown to induce an imbalance in cytotoxicity cellular antioxidant/oxidant status, and DNA damage in mammalian cell lines. This study was designed to evaluate the protective effect of ascorbic acid (Asc) against the cytotoxicity, reactive oxygen species (ROS) production, genotoxicity and epigenetic changes induced by 3,5‐DMAP in AA8 Chinese Hamster Ovary (CHO) cells. In different cellular fractions, 3,5‐DMAP caused alterations in the enzyme activities orchestrating a cellular antioxidant balance, decreases in reduced glutathione levels and a cellular redox ratio as well as increases in lipid peroxidation and protein oxidation. We also suggest that the cellular stress caused by this particular alkylaniline leads to both genetic (Aprt mutagenesis) and epigenetic changes in histones 3 and 4 (H3 and H4). This may further cause molecular events triggering different pathological conditions and eventually cancer. In both cytoplasm and nucleus, Asc provided increases in 3,5‐DMAP‐reduced glutathione levels and cellular redox ratio and decreases in the lipid peroxidation and protein oxidation. Asc was also found to be protective against the genotoxic and epigenetic effects initiated by 3,5‐DMAP. In addition, Asc supplied protection against the cell cycle (G1 phase) arrest induced by this particular alkylaniline metabolite.
      PubDate: 2014-09-01T06:51:00.294209-05:
      DOI: 10.1002/jat.3046
       
  • In utero and early childhood exposure to arsenic decreases lung function
           in children
    • Authors: Rogelio Recio‐Vega; Tania Gonzalez‐Cortes, Edgar Olivas‐Calderon, R. Clark Lantz, A. Jay Gandolfi, Cesar Gonzalez‐De Alba
      Abstract: The lung is a target organ for adverse health outcomes following exposure to As. Several studies have reported a high prevalence of respiratory symptoms and diseases in subjects highly exposed to As through drinking water; however, most studies to date has been performed in exposed adults, with little information on respiratory effects in children. The objective of the study was to evaluate the association between urinary levels of As and its metabolites with lung function in children exposed in utero and in early childhood to high As levels through drinking water. A total of 358 healthy children were included in our study. Individual exposure was assessed based on urinary concentration of inorganic As. Lung function was assessed by spirometry. Participants were exposed since pregnancy until early childhood to an average water As concentration of 152.13 µg l–1. The mean urinary As level registered in the studied subjects was 141.2 µg l–1 and only 16.7% had a urinary concentration below the national concern level. Forced vital capacity was significantly decreased in the studied population and it was negatively associated with the percentage of inorganic As. More than 57% of the subjects had a restrictive spirometric pattern. The urinary As level was higher in those children with restrictive lung patterns when compared with the levels registered in subjects with normal spirometric patterns. Exposure to As through drinking water during in utero and early life was associated with a decrease in forced vital capacity and with a restrictive spirometric pattern in the children evaluated. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-15T08:01:31.55848-05:0
      DOI: 10.1002/jat.3023
       
  • Development of haemostatic decontaminants for the treatment of wounds
           contaminated with chemical warfare agents. 1: Evaluation of in vitro
           clotting efficacy in the presence of certain contaminants
    • Authors: Charlotte A. Hall; Helen L. Lydon, Christopher H. Dalton, J. K. Chipman, John S. Graham, Robert P. Chilcott
      Abstract: The treatment of penetrating, haemorrhaging injuries sustained within a hazardous environment may be complicated by contamination with toxic chemicals. There are currently no specific medical countermeasures for such injuries. Haemostats with an absorbent mechanism of action have the potential to simultaneously stop bleeding and decontaminate wounds. However, a primary requirement of a ‘haemostatic decontaminant’ is the retention of clotting function in the presence of chemical contaminants. Thus, the aim of this study was to investigate the haemostatic efficacy of seven commercially available haemostats in the presence of toxic chemicals (soman, VX, sulphur mustard, petrol, aviation fuel and motor oil). Clot viscosity was assessed ex vivo using thrombelastography following treatment of pig blood with: (i) toxic chemical; (ii) haemostat; or (iii) haemostat in combination with toxic chemical. Several contaminants (VX, petrol and GD) were found to be pro‐haemostatic and none had an adverse effect on the rate with which the test products attained haemostasis. However, the total clot strength for blood treated with certain haemostats in the presence of sulphur mustard, soman and petrol was significantly decreased. Three test products failed to demonstrate haemostatic function in this ex vivo (thrombelastography) model; this was tentatively ascribed to the products achieving haemostasis through a tamponade mechanism of action, which can only be replicated using in vivo models. Overall, this study has identified a number of commercial products that may have potential as haemostatic decontaminants and warrant further investigation to establish their decontaminant efficacy. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-15T07:46:27.370339-05:
      DOI: 10.1002/jat.3019
       
  • Systemic drugs inducing non‐immediate cutaneous adverse reactions
           and contact sensitizers evoke similar responses in THP‐1 cells
    • Authors: Margarida Gonçalo; João Martins, Ana Silva, Bruno Neves, Américo Figueiredo, Teresa Cruz, Celeste Lopes
      Abstract: Contact sensitizers induce phenotypic and functional changes in dendritic cells (DC) that enhance their antigen‐presenting capacity and, ultimately, modulate the T cell response. To evaluate if there is a similar effect of drugs causing T‐cell‐mediated cutaneous adverse drug reactions (CADR), we studied the in vitro effect of drugs on THP‐1 cells, a cell line widely used to evaluate the early molecular and cellular events triggered by contact sensitizers. The effect of allopurinol, oxypurinol, ampicillin, amoxicillin, carbamazepine and sodium valproate, at EC30 concentrations, was evaluated on p38 MAPK activation, by Western Blot, and on the expression of genes coding for DC maturation markers, pro‐inflammatory cytokine/chemokines and hemeoxygenase 1 (HMOX1), by real‐time RT‐PCR. Results were compared with lipopolysaccharide (LPS), a DC maturation stimulus, and the strong contact sensitizer, 1‐fluoro‐2,4‐dinitrobenzene (DNFB). All drugs studied significantly upregulated HMOX1 gene transcription and all, except the anticonvulsants, also upregulated IL8. Allopurinol and oxypurinol showed the most intense effect, in a magnitude similar to DNFB and superior to betalactams. Transcription of CD40, IL12B and CXCL10 genes by drugs was more irregular. Moreover, like DNFB, all drugs activated p38 MAPK, although significantly only for oxypurinol. Like contact sensitizers, drugs that cause non‐immediate CADR activate THP‐1 cells in vitro, using different signalling pathways and affecting gene transcription with an intensity that may reflect the frequency and severity of the CADR they cause. Direct activation of antigen‐presenting DC by systemic drugs may be an important early step in the pathophysiology of non‐immediate CADR. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:46:53.569007-05:
      DOI: 10.1002/jat.3033
       
  • Analysis of drugs of abuse in human plasma by dispersive
           liquid–liquid microextraction and high‐performance liquid
           chromatography
    • Authors: P. Fernández; M. Regenjo, A. M. Bermejo, A. M. Fernández, R. A. Lorenzo, A. M. Carro
      Abstract: Opioids and cocaine are widely used at present, both for recreational purposes and as drugs of abuse. This raises the need to develop new analytical methods specifically designed for the simultaneous detection of several drugs of abuse in biological samples. In this work, dispersive liquid–liquid microextraction (DLLME) was assessed as a new sample treatment for the simultaneous extraction of morphine (MOR), 6‐acetylmorphine (6AM), cocaine (COC), benzoylecgonine (BZE) and methadone (MET) from human plasma. Preliminary assays were done before developing an experimental design based on a Uniform Network Doehlert which allowed the optimum extraction conditions to be identified, namely: a volume of extractant solvent (chloroform) and dispersant solvent (acetonitrile) of 220 µl and 3.2 ml, respectively; 0.2 g of NaCl as a salting‐out additive; pH 10.6 and ultrasound stirring for 3.5 min. The resulting extracts were analyzed by high‐performance liquid chromatography with photodiode array detection (HPLC‐PDA), using an XBridge® RP18 column (250 × 4.6 mm i.d., 5 µm particle size). Calibration graphs were linear over the concentration range 0.1–10 µg ml–1, and detection limits ranged from 13.9 to 28.5 ng ml–1. Precision calculated at three different concentration levels in plasma was included in the range 0.1–6.8% RSD. Recoveries of the five drugs were all higher than 84% on average. Finally the proposed method was successfully applied to 22 plasma samples from heroin, cocaine and/or methadone users, and the most frequently detected drug was benzoylecgonine, followed by methadone, cocaine and morphine. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:42:27.742954-05:
      DOI: 10.1002/jat.3035
       
  • The relationship between chemical‐induced kidney weight increases
           and kidney histopathology in rats
    • Authors: Evisabel A. Craig; Zhongyu Yan, Q. Jay Zhao
      Abstract: The kidney is a major site of chemical excretion, which results in its propensity to exhibit chemically‐induced toxicological effects at a higher rate than most other organs. Although the kidneys are often weighed in animal toxicity studies, the manner in which these kidney weight measurements are interpreted and the value of this information in predicting renal damage remains controversial. In this study we sought to determine whether a relationship exists between chemically‐induced kidney weight changes and renal histopathological alterations. We also examined the relative utility of absolute and relative (kidney‐to‐body weight ratio) kidney weight in the prediction of renal toxicity. For this, data extracted from oral chemical exposure studies in rats performed by the National Toxicology Program were qualitatively and quantitatively evaluated. Our analysis showed a statistically significant correlation between absolute, but not relative, kidney weight and renal histopathology in chemically‐treated rats. This positive correlation between absolute kidney weight and histopathology was observed even with compounds that statistically decreased terminal body weight. Also, changes in absolute kidney weight, which occurred at subchronic exposures, were able to predict the presence or absence of kidney histopathology at both subchronic and chronic exposures. Furthermore, most increases in absolute kidney weight reaching statistical significance (irrespective of the magnitude of change) were found to be relevant for the prediction of histopathological changes. Hence, our findings demonstrate that the evaluation of absolute kidney weight is a useful method for identifying potential renal toxicants. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:42:22.00032-05:0
      DOI: 10.1002/jat.3036
       
  • Protective role of L‐ascorbic acid, N‐acetylcysteine and
           apocynin on neomycin‐induced hair cell loss in Zebrafish
    • Authors: Chia‐Yen Wu; Han‐Jung Lee, Chi‐Fang Liu, Mallikarjuna Korivi, Hwei‐Hsien Chen, Ming‐Huan Chan
      Abstract: Hair cells are highly sensitive to environmental insults and other therapeutic drugs. The adverse effects of drugs such as aminoglycosides can cause hair cell death and lead to hearing loss and imbalance. The objective of the present study was to evaluate the protective activity of L‐ascorbic acid, N‐acetylcysteine (NAC) and apocynin on neomycin‐induced hair cell damage in zebrafish (Danio rerio) larvae at 5 days post fertilization (dpf). Results showed that the loss of hair cells within the neuromasts of the lateral lines after neomycin exposure was evidenced by a significantly lower number of neuromasts labeled with fluorescent dye FM1‐43FX observed under a microscope. Co‐administration with L‐ascorbic acid, NAC and apocynin protected neomycin‐induced hair cell loss within the neuromasts. Moreover, these three compounds reduced the production of reactive oxygen species (ROS) in neuromasts exposed to neomycin, indicating that their antioxidant action is involved. In contrast, the neuromasts were labeled with specific fluorescent dye Texas‐red conjugated with neomycin to detect neomycin uptake. Interestingly, the uptake of neomycin into hair cells was not influenced by these three antioxidant compounds. These data imply that prevention of hair cell damage against neomycin by L‐ascorbic acid, NAC and apocynin might be associated with inhibition of excessive ROS production, but not related to modulating neomycin uptake. Our findings conclude that L‐ascorbic acid, NAC and apocynin could be used as therapeutic drugs to protect aminoglycoside‐induced listening impairment after further confirmatory studies. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:42:16.707002-05:
      DOI: 10.1002/jat.3043
       
  • Plasma miR‐208 as a useful biomarker for drug‐induced
           cardiotoxicity in rats
    • Authors: Yoko Nishimura; Chiaki Kondo, Yuji Morikawa, Yutaka Tonomura, Mikinori Torii, Jyoji Yamate, Takeki Uehara
      Abstract: Cardiotoxicity is one of the major safety concerns in drug development. Therefore, detecting and monitoring cardiotoxicity throughout preclinical and clinical studies is important for pharmaceutical companies. The present study was conducted in order to explore a plasma miRNA biomarker for cardiotoxicity in rats. As organ specificity is an important factor for a biomarker, we analyzed the miRNA microarray dataset in 55 organs/tissues in normal male rats. Based on this analysis, 5 miRNAs consisting of miR‐208 (heart‐specific), miR‐1, miR‐133a, miR‐133b (heart and skeletal muscle‐specific) and miR‐206 (skeletal muscle‐specific) were selected. Next, we evaluated the usefulness of those 5 miRNAs as circulating biomarkers in rats administered with single‐dose isoproterenol or doxorubicin. Plasma miR‐208 was consistently increased through 24 h after dosing in rats administered with isoproterenol, whereas plasma concentrations of cardiac troponin (cTn) showed transient elevation. In contrast, the plasma levels of miR‐1, miR‐133a, miR‐133a and miR‐206 were elevated after treatment with doxorubicin, probably as a result of skeletal muscle toxicity. Additionally, the plasma miR‐208 level was elevated even after repeat‐dose administration (once daily for 7 days) of isoproterenol under which the pathological condition proceeded to the sub‐chronic phase such as fibrosis. Thus, our data suggest that miR‐208 is a promising plasma biomarker for cardiotoxicity in rats. Monitoring of plasma miR‐208 levels in rats may lead to more accurate evaluation of cardiotoxicity in preclinical studies. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:03:05.796088-05:
      DOI: 10.1002/jat.3044
       
  • In vitro evaluation of the effects of perfluorooctanesulfonic acid (PFOS)
           and perfluorooctanoic acid (PFOA) on IL‐2 production in human
           T‐cells
    • Authors: Kristin Midgett; Margie M. Peden‐Adams, Gary S. Gilkeson, Diane L. Kamen
      Abstract: Perfluorinated compounds, such as perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA), have been shown to alter various immune functions suggesting they are immunotoxic. This study assessed the effects of PFOS and PFOA on interleukin (IL)‐2 production in the human Jurkat T‐cell line and PFOS in healthy human primary T cells. Jurkat cells were stimulated with phytohemagglutinin (PHA)/phorbol myristate acetate (PMA), anti CD‐3/anti CD‐28, or anti CD‐3, and dosed with 0, 0.05, 0.1, 0.5, 1, 5, 10, 50, 75, or 100 µg ml−1 PFOS or 0, 0.005, 0.01, 0.05, 0.1, 0.5, 1, 5, or 10 µg ml−1 PFOA. Jurkat cells stimulated with PHA/PMA or anti CD‐3 exhibited decreased IL‐2 production beginning at 50 µg PFOS ml−1 and 5 µg PFOS ml−1 respectively, but stimulation with anti‐CD3/anti‐CD28 resulted in no changes compared with the control. Addition of the PPAR‐alpha antagonist GW6471 to PFOS‐dosed cells stimulated with PHA/PMA resulted in decreases in IL‐2 production starting at 50 µg PFOS ml−1, which suggests PFOS affected T‐cell IL‐2 production via PPAR‐alpha‐independent mechanisms. Exposure to PFOA, PFOA + GW6471, or PFOS + PFOA in Jurkat cells resulted in no significant differences in IL‐2 production. In vitro dosing studies using healthy primary human CD4+ T cells were consistent with the Jurkat results. These data demonstrated that PFOA did not impact IL‐2 production, but PFOS suppressed IL‐2 production in both a human cell line and human primary cells at dose levels within the high end of the human exposure range. A decrease in IL‐2 production is characteristic of autoimmune diseases such as systemic lupus erythematosus and should be further investigated. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-23T08:44:08.47006-05:0
      DOI: 10.1002/jat.3037
       
  • Characterization of Oryzias latipes glucocorticoid receptors and their
           unique response to progestins
    • Authors: Shinichi Miyagawa; Anke Lange, Saki Tohyama, Yukiko Ogino, Takeshi Mizutani, Tohru Kobayashi, Norihisa Tatarazako, Charles R. Tyler, Taisen Iguchi
      Abstract: Various receptor bioassays, including estrogens, androgens and thyroid hormones, have been developed and applied successfully for assessing hormone function in a wide range of animal species, including fish. In fish, corticosteroids play a pivotal role in physiology as they do in mammals, but far less is known about the corticosteroid receptor system in fish compared with in mammals. Here we established a transient transactivation assay using the Japanese medaka, Oryzias latipes, glucocorticoid receptors (olGRs) and mineralocorticoid receptor to analyse their functional properties in a fish. We found that olGR2 was highly responsive to glucocorticoids, similar to the human GR, whereas the olGR1 subtype was minimally responsive. Thus, olGR2 most likely mediates glucocorticoid signaling in medaka. We further tested crosstalk between GRs and other steroid hormones, and found that progestins could activate or inactivate olGR2‐mediating transcription, depending on the presence or absence of cortisol. The transactivation assays developed for medaka GRs provide tools to gain useful insights into corticosteroid signaling in fish and for in vitro screening of environmental substances activating GRs. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-23T08:42:52.17801-05:0
      DOI: 10.1002/jat.3020
       
  • Cytotoxic and apoptotic activities of the plancitoxin I from the venom of
           crown‐of‐thorns starfish (Acanthaster planci) on A375.S2 cells
           
    • Authors: Chi‐Chiu Lee; Hernyi Justin Hsieh, Deng‐Fwu Hwang
      Abstract: This study reports on a cytotoxic toxin derived from the venom of the crown‐of‐thorns starfish Acanthaster planci (CAV). The protein toxin was isolated through both ion‐exchange and gel‐filtration chromatography, and characterized by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) and mass spectrum analyzes. The CAV was identified as plancitoxin I protein. The mechanistic role of the CAV toxin was explored in human malignant melanoma A375.S2 cell death. The results indicated that after incubation with CAV toxin, cells significantly decreased in A375.S2 cell viability and increased in the lactate dehydrogenase (LDH) level in a dose‐dependent manner. The assays indicated that CAV toxin promoted reactive oxygen species (ROS) production, induced nitric oxide (NO) formation, lost mitochondrial membrane potential (ΔΨm) and induced inter‐nucleosomal DNA fragmentation in A375.S2 cells. The molecular cytotoxicity of the CAV toxin was tested through evaluation of the apoptosis/necrosis ratio by double staining with annexin V‐FITC and a propidium iodide (PI) assay. The results suggested that CAV toxin induced a cytotoxic effect in A375.S2 cells via the apoptotic procedure, and may be associated with the regulation of the p38 pathways. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-21T09:24:19.994647-05:
      DOI: 10.1002/jat.3034
       
  • Elevated levels of antibodies against xenobiotics in a subgroup of healthy
           subjects
    • Authors: Aristo Vojdani; Datis Kharrazian, Partha Sarathi Mukherjee
      Abstract: In spite of numerous research efforts, the exact etiology of autoimmune diseases remains largely unknown. Genetics and environmental factors, including xenobiotics, are believed to be involved in the induction of autoimmune disease. Some environmental chemicals, acting as haptens, can bind to a high‐molecular‐weight carrier protein such as human serum albumin (HSA), causing the immune system to misidentify self‐tissue as an invader and launch an immune response against it, leading to autoimmunity. This study aimed to examine the percentage of blood samples from healthy donors in which chemical agents mounted immune challenges and produced antibodies against HSA‐bound chemicals. The levels of specific antibodies against 12 different chemicals bound to HSA were measured by ELISA in serum from 400 blood donors. We found that 10% (IgG) and 17% (IgM) of tested individuals showed significant antibody elevation against aflatoxin‐HSA adduct. The percentage of elevation against the other 11 chemicals ranged from 8% to 22% (IgG) and 13% to 18% (IgM). Performance of serial dilution and inhibition of the chemical–antibody reaction by specific antigens but not by non‐specific antigens were indicative of the specificity of these antibodies. Although we lack information about chemical exposure in the tested individuals, detection of antibodies against various protein adducts may indicate chronic exposure to these chemical haptens in about 20% of the tested individuals. Currently the pathological significance of these antibodies in human blood is still unclear, and this protein adduct formation could be one of the mechanisms by which environmental chemicals induce autoimmune reactivity in a significant percentage of the population. Copyright © 2014. The
      Authors . Journal of Applied Toxicology Published by John Wiley & Sons Ltd.
      PubDate: 2014-07-18T04:30:19.876942-05:
      DOI: 10.1002/jat.3031
       
  • Reactive oxygen species‐dependent JNK downregulated
           olaquindox‐induced autophagy in HepG2 cells
    • Authors: Dongxu Zhao; Congcong Wang, Shusheng Tang, Chaoming Zhang, Shen Zhang, Yan Zhou, Xilong Xiao
      Abstract: Autophagy plays an important role in response to intracellular and extracellular stress to sustain cell survival. However, dysregulated or excessive autophagy may lead to cell death, known as “type II programmed cell death,” and it is closely associated with apoptosis. In our previous study, we proposed that olaquindox induced apoptosis of HepG2 cells through a caspase‐9 dependent mitochondrial pathway. In this study, we investigated autophagy induced by olaquindox and explored the crosstalk between apoptosis and autophagy in olaquindox‐treated HepG2 cells. Olaquindox‐induced autophagy was demonstrated by the accumulation of monodansylcadervarine, as well as elevated expression of autophagy‐related MAP‐LC3 and Beclin 1 proteins. The autophagy inhibitor 3‐methyladenine significantly increased the apoptotic rate induced by olaquindox, which was correlated with increased ratio of Bax/Bcl‐2. The further studies showed that olaquindox increased the levels of reactive oxygen species (ROS), and antioxidant N‐acetyl‐L‐cysteine (NAC) effectively blocked the accumulation of ROS but failed to block autophagy. Moreover, olaquindox induced the activation of c‐Jun N‐terminal protein kinase (JNK), and JNK inhibitor SP600125 failed to block autophagy. Instead, olaquindox‐induced autophagy was enhanced by NAC or SP600125. Meanwhile, JNK activation was remarkably blocked by NAC, indicating that ROS may be the upstream signaling molecules of JNK activation and involved in the negative regulation of olaquindox‐induced autophagy. These results suggest that olaquindox induces autophagy in HepG2 cells and that olaquindox‐induced apoptosis can be enhanced by 3‐methyladenine. Olaquindox‐induced autophagy in HepG2 cells is upregulated by Beclin 1 but downregulated by ROS‐dependent JNK. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-18T04:30:06.618358-05:
      DOI: 10.1002/jat.3022
       
  • Inhibitory effect of apocynin on methylglyoxal‐mediated glycation in
           osteoblastic MC3T3‐E1 cells
    • Authors: Kwang Sik Suh; Sang Youl Rhee, Young Seol Kim, Eun Mi. Choi
      Abstract: Methylglyoxal (MG), a highly reactive metabolite of hyperglycemia, can enhance protein glycation, oxidative stress or inflammation. The present study investigated the effects of apocynin on the mechanisms associated with MG toxicity in osteoblastic MC3T3‐E1 cells. Pretreatment of MC3T3‐E1 cells with apocynin prevented the MG‐induced protein glycation and formation of intracellular reactive oxygen species and mitochondrial superoxide in MC3T3‐E1 cells. In addition, apocynin increased glutathione levels and restored the activity of glyoxalase I inhibited by MG. These findings suggest that apocynin provide a protective action against MG‐induced cell damage by reducing oxidative stress and by increasing the MG detoxification system. Apocynin treatment decreased the levels of proinflammatory cytokines such as tumor necrosis factor‐α and interleukin‐6 induced by MG. Additionally, the nitric oxide level reduced by MG was significantly increased by apocynin. These findings indicate that apocynin might exert its therapeutic effects via upregulation of glyoxalase system and antioxidant activity. Taken together, apocynin may prove to be an effective treatment for diabetic osteopathy. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-18T04:30:03.532821-05:
      DOI: 10.1002/jat.3016
       
  • Renal cells exposed to cadmium in vitro and in vivo: normalizing gene
           expression data
    • Authors: A. R. Nair; K. Smeets, E. Keunen, W.‐K. Lee, F. Thévenod, E. Van Kerkhove, A. Cuypers
      Abstract: Cadmium (Cd) is a toxic metal with a long half‐life in biological systems. This half‐life is partly as a result of metallothioneins (MTs), metal‐binding proteins with a high affinity for Cd. The high retention properties of the kidneys reside in proximal tubular cells that possess transport mechanisms for Cd‐MT uptake, ultimately leading to more Cd accumulation. Researchers have studied MT–metal interactions using various techniques including quantitative real‐time PCR (qPCR), an efficient tool for quantifying gene expression. Often a poor choice of reference genes, which is represented by their instability and condition dependency, leads to inefficient normalization of gene expression data and misinterpretations. This study demonstrates the importance of an efficient normalization strategy in toxicological research. A selection of stable reference genes was proposed in order to acquire reliable and reproducible gene quantification under metal stress using MT expression as an example. Moreover, in vitro and in vivo setups were compared to identify the influence of toxicological compounds in function of the experimental design. This study shows that glyceraldehyde‐3‐phosphate dehydrogenase (Gapdh), tyrosine monooxygenase/tryptophan5‐monooxygenase activation‐protein, zeta polypeptide (Ywhaz) and beta‐actin (Actb) are the most stable reference genes in a kidney proximal tubular cell line exposed to moderate and high Cd concentrations, applied as CdCl2. A slightly different sequence in reference gene stability was found in renal cells isolated from rats in vivo exposed to Cd. It was further shown that three reference genes are required for efficient normalization in this experimental setup. This study demonstrates the importance of an efficient normalization strategy in toxicological research. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-18T04:29:56.735907-05:
      DOI: 10.1002/jat.3047
       
  • Toxicity of new emerging pollutant tris‐(2,3‐dibromopropyl)
           isocyanurate on BALB/c mice
    • Authors: Juan Li; Xu Zhang, Jieqing Bao, Yuchen Liu, Junfeng Li, Jia Li, Yong Liang, Jie Zhang, Aiqian Zhang
      Abstract: The emerging heterocyclic brominated flame retardant tris‐(2,3‐dibromopropyl) isocyanurate (TBC), widely used in reinforced plastics, has demonstrated toxicity to fish. However, little is known about its toxicity in rodents. This study aims to determine the effect of TBC on growth, biochemical parameters in serum, organs and related gene expression of both male and female BALB/c mice after gastro‐gavage administration of 0, 2, 10 and 50 mg kg−1 TBC for 28 days. Results indicated that exposure to TBC had no effects on basic growth and food intake of mice, but significantly increased serum alanine aminotransferase levels in male mice. Histopathological analyses showed that focal necrosis (2, 10 and 50 mg kg−1 TBC‐exposed groups) and ballooning degeneration (10 and 50 mg kg−1 TBC‐exposed groups) were found in mouse liver, whereas transmission electron microscopy revealed dose‐dependent hepatocyte apoptosis, mitochondrial degeneration and endoplasmic reticulum dilation. Histopathological and ultrastructural assessments in the lung showed dose‐dependent hyperplasia of pulmonary alveolar epithelium, bronchial congestion, infiltration of inflammatory cells and mitochondrial swelling following TBC exposure. Our results also indicated that mitochondria are one of the major target cytoplasmic organelles for TBC, suggesting that damage in mitochondria is one of the pathways that led to toxic effects in the liver and lung of TBC‐treated groups. Moreover, TBC effectively activated the gene expression of p53 in mice liver. Our findings provide strong evidence that TBC induces significant toxicity in mice organs, especially in liver and lung, which play vital roles in detoxification and gas exchange, respectively. This research will contribute to characterize the toxic effects of TBC, which was introduced as one of the candidates for brominated flame retardant replacement. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-17T23:01:48.126209-05:
      DOI: 10.1002/jat.3026
       
  • Toxicity profiles and solvent–toxicant interference in the planarian
           Schmidtea mediterranea after dimethylsulfoxide (DMSO) exposure
    • Authors: An‐Sofie Stevens; Nicky Pirotte, Michelle Plusquin, Maxime Willems, Thomas Neyens, Tom Artois, Karen Smeets
      Abstract: To investigate hydrophobic test compounds in toxicological studies, solvents like dimethylsulfoxide (DMSO) are inevitable. However, using these solvents, the interpretation of test compound‐induced responses can be biased. DMSO concentration guidelines are available, but are mostly based on acute exposures involving one specific toxicity endpoint. Hence, to avoid solvent–toxicant interference, we use multiple chronic test endpoints for additional interpretation of DMSO concentrations and propose a statistical model to assess possible synergistic, antagonistic or additive effects of test compounds and their solvents. In this study, the effects of both short‐ (1 day) and long‐term (2 weeks) exposures to low DMSO concentrations (up to 1000 µl l−1) were studied in the planarian Schmidtea mediterranea. We measured different biological levels in both fully developed and developing animals. In a long‐term exposure set‐up, a concentration of 500 µl l−1 DMSO interfered with processes on different biological levels, e.g. behaviour, stem cell proliferation and gene expression profiles. After short exposure times, 500 µl l−1 DMSO only affected motility, whereas the most significant changes on different parameters were observed at a concentration of 1000 µl l−1 DMSO. As small sensitivity differences exist between biological levels and developmental stages, we advise the use of this solvent in concentrations below 500 µl l−1 in this organism. In the second part of our study, we propose a statistical approach to account for solvent–toxicant interactions and discuss full‐scale solvent toxicity studies. In conclusion, we reassessed DMSO concentration limits for different experimental endpoints in the planarian S. mediterranea. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-06-25T22:54:19.05537-05:0
      DOI: 10.1002/jat.3011
       
  • Vascular endothelial growth factor mRNA levels as a biomarker for
           short‐term N‐butyl‐N‐(4‐hydroxybutyl)
           nitrosamine‐induced rat bladder carcinogenesis bioassay
    • Authors: Shin Wakui; Tomoko Mutou, Hiroyuki Takahashi, Masahiro Ikegami, Hideki Wanibuchi, Shoji Fukushima
      Abstract: Generically, carcinogenic effects of chemicals in bladder carcinogenesis are judged by induction of papillary or nodular (PN) hyperplasia in rats given N‐butyl‐N‐(4‐hydroxybutyl) nitrosamine (BBN) for 4 weeks and the test chemical for 22–28 weeks. However, upregulation of vascular endothelial growth factor (VEGF) begins early in rat BBN bladder carcinogenesis. To establish a short‐term rat bladder carcinogenic bioassay, we analyzed the correlations between VEGF, VEGF mRNA and bladder lesions inductions at 10 and 26 weeks after BBN treatment. Six‐week‐old male Wistar (slc) rats were given 0.05% BBN for 4, 10 or 26 weeks. To avoid individual rat bias, the bladders were investigated by partial cystectomy at 10 weeks and total cystectomy at 26 weeks. After induction, PN hyperplasia and carcinoma in rats increased with the length of BBN treatment and immunohistochemical VEGF expression also increased following carcinogenesis, but the immunoreactivity of individual lesions was quite variable. Moreover, induction of PN hyperplasia at 10 weeks’ BBN treatment was not significantly correlated with that at 26 weeks' treatment; thus, it was not possible to predict the carcinogenic effect due to the induction of PN hyperplasia at 26 weeks' BBN treatment by that at 10 weeks' treatment. However, VEGF mRNA levels of rat bladders at 10 weeks' BBN treatment revealed a strong significant correlation with the incidence of bladder lesions at 26 weeks' treatment. Here, we suggest that quantitative VEGF mRNA levels are a good biomarker for a short‐term BBN‐induced bioassay for rat bladder carcinogenesis. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-28T09:25:42.146279-05:
      DOI: 10.1002/jat.3021
       
  • Developmental toxicity assay using high content screening of zebrafish
           embryos
    • Authors: Susan Lantz‐McPeak; Xiaoqing Guo, Elvis Cuevas, Melanie Dumas, Glenn D. Newport, Syed F. Ali, Merle G. Paule, Jyotshna Kanungo
      Abstract: Typically, time‐consuming standard toxicological assays using the zebrafish (Danio rerio) embryo model evaluate mortality and teratogenicity after exposure during the first 2 days post‐fertilization. Here we describe an automated image‐based high content screening (HCS) assay to identify the teratogenic/embryotoxic potential of compounds in zebrafish embryos in vivo. Automated image acquisition was performed using a high content microscope system. Further automated analysis of embryo length, as a statistically quantifiable endpoint of toxicity, was performed on images post‐acquisition. The biological effects of ethanol, nicotine, ketamine, caffeine, dimethyl sulfoxide and temperature on zebrafish embryos were assessed. This automated developmental toxicity assay, based on a growth‐retardation endpoint should be suitable for evaluating the effects of potential teratogens and developmental toxicants in a high throughput manner. This approach can significantly expedite the screening of potential teratogens and developmental toxicants, thereby improving the current risk assessment process by decreasing analysis time and required resources. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.
      PubDate: 2014-05-28T09:20:58.316539-05:
      DOI: 10.1002/jat.3029
       
  • Cardiotoxicity evaluation of anthracyclines in zebrafish (Danio rerio)
    • Authors: Ying Han; Jing‐pu Zhang, Jian‐qin Qian, Chang‐qin Hu
      Abstract: Drug‐induced cardiotoxicity is a leading factor for drug withdrawals, and limits drug efficacy and clinical use. Therefore, new alternative animal models and methods for drug safety evaluation have been given great attention. Anthracyclines (ANTs) are widely prescribed anticancer agents that have a cumulative dose relationship with cardiotoxicity. We performed experiments to study the toxicity of ANTs in early developing zebrafish embryos, especially their effects on the heart. LC50 values for daunorubicin, pirarubicin, doxorubicin (DOX), epirubicin and DOX‐liposome at 72 h post‐fertilization were 122.7 μM, 111.9 μM, 31.2 μM, 108.3 μM and 55.8 μM, respectively. At the same time, zebrafish embryos were exposed to ANTs in three exposure stages and induced incomplete looping of the heart tube, pericardia edema and bradycardia in a dose‐dependent manner, eventually leading to death. DOX caused the greatest heart defects in the treatment stages and its liposome reduced the effects on the heart, while daunorubicin produced the least toxicity. Genes and proteins related to heart development were also identified to be sensitive to ANT exposure and downregulated by ANTs. It revealed ANTs could disturb the heart formation and development. ANTs induced cardiotoxicity in zebrafish has similar effects in mammalian models, indicating that zebrafish may have a potential value for assessment of drug‐induced developmental cardiotoxicity. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-22T16:22:52.444139-05:
      DOI: 10.1002/jat.3007
       
  • A trivalent approach for determining in vitro toxicology: Examination of
           oxime K027
    • Authors: Adriana Prado; Georg A. Petroianu, Dietrich E. Lorke, Jeremy W. Chambers
      Abstract: Unforeseen toxic effects contribute to compound attrition during preclinical evaluation and clinical trials. Consequently, there is a need to correlate in vitro toxicity to in vivo and clinical outcomes quickly and effectively. We propose an expedited evaluation of physiological parameters in vitro that will improve the ability to predict in vivo toxicity of potential therapeutics. By monitoring metabolism, mitochondrial physiology and cell viability, our approach provides insight to the extent of drug toxicity in vitro. To implement our approach, we used human hepatocellular carcinoma cells (HepG2) and neuroblastoma cells (SH‐SY5Y) to monitor hepato‐ and neurotoxicity of the experimental oxime K027. We utilized a trivalent approach to measure metabolism, mitochondrial stress and induction of apoptosis in 96‐well formats. Any change in these three areas may suggest drug‐induced toxicity in vivo. K027 and pralidoxime, an oxime currently in clinical use, had no effect on glycolysis or oxygen consumption in HepG2 and SH‐SY5Y cells. Similarly, these oximes did not induce oxidant generation nor alter mitochondrial membrane potential. Further, K027 and pralidoxime failed to activate effector caspases, and these oximes did not alter viability. The chemotherapeutic agent, docetaxel, negatively affected metabolism, mitochondrial physiology and viability. Our studies present a streamlined high‐throughput trivalent approach for predicting toxicity in vitro, and this approach reveals that K027 has no measurable hepatotoxicity or neurotoxicity in vitro, which correlates with their in vivo data. This approach could eliminate toxic drugs from consideration for in vivo preclinical evaluation faster than existing toxicity prediction panels and ultimately prevent unnecessary experimentation. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-22T16:22:49.472421-05:
      DOI: 10.1002/jat.3013
       
  • Synergistic cytotoxicity and DNA strand breaks in cells and plasmid DNA
           exposed to uranyl acetate and ultraviolet radiation
    • Authors: Janice Wilson; Mary C. Zuniga, Filbert Yazzie, Diane M. Stearns
      Abstract: Depleted uranium (DU) has a chemical toxicity that is independent of its radioactivity. The purpose of this study was to explore the photoactivation of uranyl ion by ultraviolet (UV) radiation as a chemical mechanism of uranium genotoxicity. The ability of UVB (302 nm) and UVA (368 nm) radiation to photoactivate uranyl ion to produce single strand breaks was measured in pBR322 plasmid DNA, and the presence of adducts and apurinic/apyrimidinic sites that could be converted to single strand breaks by heat and piperidine was analyzed. Results showed that DNA lesions in plasmid DNA exposed to UVB‐ or UVA‐activated DU were only slightly heat reactive, but were piperidine sensitive. The cytotoxicity of UVB‐activated uranyl ion was measured in repair‐proficient and repair‐deficient Chinese hamster ovary cells and human keratinocyte HaCaT cells. The cytotoxicity of co‐exposures of uranyl ion and UVB radiation was dependent on the order of exposure and was greater than co‐exposures of arsenite and UVB radiation. Uranyl ion and UVB radiation were synergistically cytotoxic in cells, and cells exposed to photoactivated DU required different DNA repair pathways than cells exposed to non‐photoactivated DU. This study contributes to our understanding of the DNA lesions formed by DU, as well as their repair. Results suggest that excitation of uranyl ion by UV radiation can provide a pathway for uranyl ion to be chemically genotoxic in populations with dermal exposures to uranium and UV radiation, which would make skin an overlooked target organ for uranium exposures. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-15T09:03:01.061665-05:
      DOI: 10.1002/jat.3015
       
  • Endocrine disruption effects of p,p′‐DDE on juvenile zebrafish
    • Authors: Marta Sofia Monteiro; Maria Pavlaki, Augusto Faustino, Alexandra Rêma, Mariana Franchi, Letícia Gediel, Susana Loureiro, Inês Domingues, Jaime Rendón von Osten, Amadeu Mortágua Velho Maia Soares
      Abstract: The persistent organic pollutant p,p′‐DDE, the major metabolite of the insecticide DDT, has displayed evidence of endocrine disruption through the inhibition of androgen binding to androgen receptors in different species. Although p,p′‐DDE was continuously detected in wild fish with abnormal gonad development such as intersex, little is known about its mode of action during gonad development in fish. To elucidate the potential endocrine effects of this pollutant in zebrafish (Danio rerio), juveniles (30 days post hatch) were exposed to p,p′‐DDE during the critical window of sexual differentiation. Fish were exposed to sublethal concentrations ranging from 0.01 to 20 µg l–1 over 14 days and were maintained in control water for an additional 4 months. As core endpoints, the vitellogenin (vtg) concentration was measured at the end of exposure, and sex ratio and the gonadosomatic index were assessed 4 months after the end of exposure. An increase in vtg production in whole body homogenate was observed in fish exposed to 0.2 and 2.0 µg l–1 p,p′‐DDE. No significant differences were displayed in morphological parameters such as the gonadosomatic index of males and females or sex ratio. However, exposed females presented histopathological changes that include the reduction of the number of mature oocytes, which might impair their successful reproduction. These results demonstrate the ability of p,p′‐DDE to cause endocrine disruption in zebrafish exposed during gonad differentiation of juvenile specimens. Furthermore, vtg induction by p,p′‐DDE in juvenile zebrafish arises as a predictive marker for adverse effects of this DDT metabolite on the ovarian function of female zebrafish. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-15T08:55:11.837345-05:
      DOI: 10.1002/jat.3014
       
  • Dimethylarsinic acid (DMAv) changed the expressions of proliferative
           related factors and secretion of inflammatory cytokines in rat bladder
    • Authors: Zhang Lin; Liu Shengnan, Wang Fei, Song Yingli, Sun Qingshan, Sheng Wei, Xi. Shuhua, Sun Guifan
      Abstract: Dimethylarsinic acid (DMAV), the major urinary metabolite of inorganic arsenic, is a urinary bladder carcinogen and bladder tumor promoter in adult rats. Increased urothelial cellular proliferation has been considered as an earlier phenotype in DMAV‐induced bladder carcinogenesis. The present study examined the ultrastructural changes of bladder epithelial cells and expressions of proliferation factors, as well as the secretion of inflammatory cytokines in rats exposed to DMAV for 10 weeks by transmission electron microscopy (TEM), qRT‐PCR, immunohistochemical staining and ELISA methods. The results showed that DMAV administered in the drinking water produced urothelial cytotoxicity and ultrastructural changes in rats. PCNA, cyclin D1 and COX‐2 mRNA expressions and immunoreactivities were elevated in bladder urothelium. In addition, 200 ppm DMAV treatment increased the transforming growth factor‐beta 1 (TGF‐β1) secretion and decreased tumor necrosis factor‐alpha (TNF)‐α level in the urine of rats. These data suggest that chronic inflammation, bladder epithelium lesions and proliferation might be the basic process of the chronic toxicity effects in DMAV‐treated rats. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-15T08:46:33.106421-05:
      DOI: 10.1002/jat.3001
       
  • Usefulness of urinary kidney injury molecule‐1 (Kim‐1) as a
           biomarker for cisplatin‐induced sub‐chronic kidney injury
    • Authors: Ken‐ichiro Nan‐ya; Masatomo Kajihara, Natsuki Kojima, Masakuni Degawa
      Abstract: We explored biomarkers suitable for monitoring sub‐chronic kidney injury using the three rat models of cisplatin (CDDP)‐induced kidney injury, which were designed to extend the current knowledge beyond the sub‐acute exposure period. In the pilot study, a single intravenous administration of 1.5 mg kg–1 CDDP to rats was confirmed to result in no histopathological changes. Subsequently, CDDP was intravenously administered to rats at a dose of 1.5 mg kg–1 for 4 days at 24‐h intervals (Experimental model 1) and for up to 10 weeks at weekly intervals (Experimental models 2 and 3), and the changes in blood and urine components, such as recently recommended urinary biomarkers (Kim‐1, clusterin and so on) and traditional blood biomarkers (blood urea nitrogen and serum creatinine), were examined together with the histopathological changes in renal tissues during the development of the kidney injury in each model. In these experimental models, a significant increase in urinary Kim‐1 was observed prior to the histopathological changes in renal tissues, and these changes were retained after the adverse histopathological changes. Significant changes in all of the other urinary biomarkers examined occurred along with the histopathological changes. In addition, the increase in urinary Kim‐1 after weekly treatment with CDDP for 4 weeks was reduced in a time‐dependent manner after cessation of the drug. The present findings indicate that urinary Kim‐1 is the most useful biomarker for CDDP‐induced rat sub‐chronic kidney injury among the biomarkers examined. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-04-16T05:43:48.985818-05:
      DOI: 10.1002/jat.2999
       
  • Evaluation of developmental toxicity using undifferentiated human
           embryonic stem cells
    • Authors: Eui‐Man Jung; Yeo‐ul Choi, Hong‐Seok Kang, Hyun Yang, Eui‐Ju Hong, Beum‐Soo An, Jun‐young Yang, Ki. Hwan Choi, Eui‐Bae Jeung
      Abstract: An embryonic stem cell test (EST) has been developed to evaluate the embryotoxic potential of chemicals with an in vitro system. In the present study, novel methods to screen toxic chemicals during the developmental process were evaluated using undifferentiated human embryonic stem (hES) cells. By using surface marker antigens (SSEA‐4, TRA‐1‐60 and TRA‐1‐81), we confirmed undifferentiated conditions of the used hES cells by immunocytochemistry. We assessed the developmental toxicity of embryotoxic chemicals, 5‐fluorouracil, indomethacin and non‐embryotoxic penicillin G in different concentrations for up to 7 days. While expressions of the surface markers were not significantly affected, the embryotoxic chemicals influenced their response to pluripotent ES cell markers, such as OCT‐4, NANOG, endothelin receptor type B (EDNRB), secreted frizzled related protein 2 (SFRP2), teratocarcinoma‐derived growth factor 1 (TDGF1), and phosphatase and tensin homolog (PTEN). Most of the pluripotent ES cell markers were down‐regulated in a dose‐dependent manner after treatment with embryotoxic chemicals. After treatment with 5‐fluorouracil, indomethacin and penicillin G, we observed a remarkable convergence in the degree of up‐regulation of development, cell cycle and apoptosis‐related genes by gene expression profiles using an Affymetrix GeneChips. Taken together, these results suggest that embryotoxic chemicals have cytotoxic effects, and modulate the expression of ES cell markers as well as development‐, cell cycle‐ and apoptosis‐related genes that have pivotal roles in undifferentiated hES cells. Therefore, we suggest that hES cells may be useful for testing the toxic effects of chemicals that could impact the embryonic developmental stage. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-04-16T05:43:46.307596-05:
      DOI: 10.1002/jat.3010
       
  • In vitro exposure of DE‐71, a penta‐PBDE mixture, on immune
           endpoints in bottlenose dolphins (Tursiops truncatus) and B6C3F1 mice
    • Authors: Jena R. Wirth; Margie M. Peden‐Adams, Natasha D. White, Gregory D. Bossart, Patricia A. Fair
      Abstract: Polybrominated diphenyl ethers (PBDEs) are an emerging contaminant of concern with low level exposures demonstrating toxicity in laboratory animals and wildlife, although immunotoxicity studies have been limited. Bottlenose dolphin peripheral blood leukocytes (PBLs) and mouse splenocytes were exposed to environmentally relevant DE‐71 (a penta‐PBDE mixture) concentrations (0–50 µg ml−1) in vitro. Natural killer (NK) cell activity and lymphocyte (B and T cell) proliferation were evaluated using the parallelogram approach for risk assessment. This study aimed to substantiate results from field studies with dolphins, assess the sensitivities between the mouse model and dolphins, and to evaluate risk using the parallelogram approach. In mouse cells, NK cell activity increased at in vitro doses 0.05, 0.5 and 25 µg DE‐71 ml−1, whereas proliferation was not modulated. In dolphin cells, NK cell activity and lymphocyte proliferation was not altered after in vitro exposure. In vitro exposure of dolphin PBLs to DE‐71 showed similar results to correlative field studies; NK cell activity in mice was more sensitive to in vitro exposure than dolphins, and the parallelogram approach showed correlation with all three endpoints to predict risk in bottlenose dolphins. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-04-07T02:26:12.183666-05:
      DOI: 10.1002/jat.3008
       
  • Impact of acute arsenic and cadmium exposure on the expression of two
           haeme oxygenase genes and other antioxidant markers in common carp
           (Cyprinus carpio)
    • Authors: Zsanett Jancsó; Edit Hermesz
      Abstract: The aim was to study the effects of cadmium (Cd) and arsenic (As) on haeme oxygenases (HOs) and other oxidative stress biomarkers, and their roles in macromolecule damage in liver and kidney of common carp (Cyprinus carpio L.). HOs play a critical role in the defence system against oxidative damage, producing biliverdin and carbon monoxide with important free radical scavenging properties. However, increased HO activity in haeme degradation may also lead to a pro‐oxidant effect through the liberation of Fe‐modifying Cd and As toxicity. The response of an organism to exposure to toxic metals is in many cases brought about by changes at the level of gene expression. In this study, the genes ho‐1 and ho‐2 of the common carp were identified, and the changes in gene expressions were analysed from the aspect of Cd and As accumulation. Both ho‐1 and ho‐2 are transcriptionally induced by Cd and As, but their inductions differ in time course, dose response and tissue specificity. The expression of ho1 was mostly affected by As, primarily in the liver (45‐fold), whereas it was enhanced with higher efficacy by Cd in the kidney (25‐fold). The cellular redox status and the damage of lipid molecules were monitored via the ratio of reduced to oxidized glutathione, the levels of H2O2 and lipid peroxidation, and the activities of superoxide dismutase (SOD) and catalase (CAT). Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-04-07T02:26:09.502028-05:
      DOI: 10.1002/jat.3000
       
  • Usefulness of optical coherence tomography to detect central serous
           chorioretinopathy in monkeys
    • Authors: Hyun‐Kyu Park; Woori Jo, Hyun‐Ji Choi, Bongcheol Kim, Gilnam Lee, Jeongbeob Seo, Suk Young Cho, Choung‐Soo Kim, Eun Kyung Choi, Jung Jin Hwang, Joo Yong Lee, Young Hee Yoon, Woo‐Chan Son
      Abstract: Many systemic drugs can induce ocular toxicity and several ocular side‐effects have been identified in clinical studies. However, it is difficult to detect ocular toxicity in preclinical studies because of the lack of appropriate evaluation methods. Optical coherence tomography (OCT) is useful because it can provide real‐time images throughout a study period, whereas histopathology only provides images of sacrificed animals. Using OCT alongside histopathology, attempts were made to find effective approaches for screening of drug‐induced ocular toxicity in monkeys. Such approaches could be used in preclinical studies prior to human trials. Six male cynomolgus monkeys (Macaca fascicularis Raffles) were orally administered one of six candidate MAPK/ERK kinase (MEK) inhibitors. Central serous chorioretinopathy, a known side‐effect of such inhibitors, was identified in four monkeys by OCT. Artifacts generated during tissue processing meant that histopathology could not detect edematous changes. Thus, OCT is a useful tool to detect ocular toxicity which cannot be detected by histopathology in preclinical studies. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-03-28T03:47:29.247172-05:
      DOI: 10.1002/jat.3009
       
  • Involvement of immune‐ and inflammatory‐related factors in
           flucloxacillin‐induced liver injury in mice
    • Authors: Shohei Takai; Satonori Higuchi, Azusa Yano, Koichi Tsuneyama, Tatsuki Fukami, Miki Nakajima, Tsuyoshi Yokoi
      Abstract: Drug‐induced liver injury (DILI) is a serious problem in pre‐clinical stages of drug development and clinical pharmacotherapy, but the pathogenesis of DILI has not been elucidated. Flucloxacillin (FLX), which is a β‐lactam antibiotic of the penicillin class that is used widely in Europe and Australia, rarely causes DILI. Clinical features suggest that FLX‐induced liver injury is caused by immune‐ and inflammatory‐related factors, but the mechanism of FLX‐induced liver injury is unknown. The purpose of this study was to elucidate the mechanisms of FLX‐induced liver injury in vivo. Plasma alanine aminotransferase, aspartate aminotransferase and total‐bilirubin levels were significantly elevated in FLX‐administered mice [1000 mg kg–1, intraperitoneally (i.p.)]. Toll‐like receptor 4 (TLR4) ligands, such as high‐mobility group box 1 (HMGB1) and S100A8/A9, were significantly increased in FLX‐administered mice, and inflammatory factors, such as interleukin (IL)‐1β, tumor necrosis factor‐alpha (TNF‐α), macrophage inflammatory protein (MIP)‐2, CXC chemokine‐ligand‐1 (CXCL1) and monocyte chemoattractant protein (MCP)‐1, were also significantly elevated. IL‐17‐related transcriptional factors and cytokines were increased, and the administration of recombinant IL‐17 (2 mg per body weight, i.p.) resulted in an exacerbation of the FLX‐induced liver injury. TLR4‐associated‐signal transduction may be involved in FLX‐induced liver injury, and IL‐17 is an exacerbating factor. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-03-20T06:29:38.953028-05:
      DOI: 10.1002/jat.3002
       
  • Molecular biomarkers of phospholipidosis in rat blood and heart after
           amiodarone treatment
    • Authors: Nicola Bocchini; Mery Giantin, Federica Crivellente, Serena Ferraresso, Ivo Faustinelli, Mauro Dacasto, Patrizia Cristofori
      Abstract: Phospholipidosis (PLD) is characterized by an intracellular accumulation of phospholipids in lysosomes and concurrent development of concentric lamellar bodies. It is induced in humans and in animals by drugs with a cationic amphiphilic structure. The purpose of the present study was to identify a set of molecular biomarkers of PLD in rat blood and heart, hypothetically applicable in preclinical screens within the drug development process. A toxicological study was set up in rats orally treated up to 11 days with 300 mg kg–1 per day–1 amiodarone (AMD). Light and transmission electron microscopy investigations were performed to confirm the presence of lamellar bodies indicative of phospholipid accumulation. The effects of AMD upon the transcriptome of these tissues were estimated using DNA microarray technology. Microarray data analysis showed that a total of 545 and 8218 genes were modulated by AMD treatment in heart and blood, respectively. Some genes implicated in the phospholipid accumulation in cells, such as phospholipase A2, showed similar alterations of gene expression. After transcriptome criteria of analysis and target selection, including also the involvement in the onset of PLD, 7 genes (Pla2g2a, Pla2g7, Gal, Il1b, Cebpb, Fcgr2b, Acer 2) were selected as candidate biomarkers of PLD in heart and blood tissues, and their potential usefulness as a sensitive screening test was screened and confirmed by quantitative Real‐Time PCR analysis. Collectively, these data underscore the importance of transcriptional profiling in drug discovery and development, and suggest blood as a surrogate tissue for possible phospholipid accumulation in cardiomyocytes. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-18T00:25:23.937611-05:
      DOI: 10.1002/jat.2992
       
  • Lipopolysaccharide exposure augments isoniazide‐induced liver injury
    • Authors: Yijing Su; Yun Zhang, Mi. Chen, Zhenzhou Jiang, Lixin Sun, Tao Wang, Luyong Zhang
      Abstract: Isoniazide (INH) is a classic antituberculosis drug associated with clinical idiosyncratic drug‐induced liver injury. It has been hypothesized that the interaction between a drug and modest inflammation results in a decreased threshold for drug toxicity. In this study, we tested the hypothesis that INH causes liver injury in rats when coadministered with lipopolysaccharide (LPS). Neither INH nor LPS alone caused liver injury. The coadministration of INH and LPS was associated with increases in serum and histopathological markers of liver injury. Tumour necrosis factor‐α expression was significantly increased in the coadministered group. The downregulation of the bile acid transporter, bile salt export pump, and multidrug resistance protein 2 at both mRNA and protein levels was observed. Furthermore, the level of Farnesoid X receptor, which regulates the bile salt export pump and multidrug resistance protein 2, were clearly decreased. These results indicate that the coadministration of nontoxic doses of LPS and INH causes liver injury; the disruption of biliary excretion is considered the primary inflammation‐related characteristic of INH‐induced hepatotoxicity. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-06T07:05:40.837044-05:
      DOI: 10.1002/jat.2979
       
  • Characterization of the biochemical effects of naphthalene on the mouse
           respiratory system using NMR‐based metabolomics
    • Authors: Jia‐Huei Hong; Wen‐Ching Lee, Yu‐Ming Hsu, Hao‐Jan Liang, Cho‐Hua Wan, Chung‐Liang Chien, Ching‐Yu Lin
      Abstract: Naphthalene is a ubiquitous environmental pollutant to which humans are exposed. Previous studies have demonstrated that naphthalene causes bronchiolar epithelial necrosis in the mouse distal airway, after parenteral administration. In this study, metabolic variations in the bronchoalveolar lavage fluid (BALF) and the lung tissues of naphthalene‐treated mice and controls were examined using nuclear magnetic resonance (NMR)‐based metabolomics to identify the toxic mechanism. Male ICR mice were treated with naphthalene [0, 50, 100 and 200 mg kg–1, intraperitoneally (i.p.)]. After 24 h, BALF and lung tissues were collected and prepared for 1H and J‐resolved (JRES) NMR analysis after principal component analysis (PCA). PCA modeling of p‐JRES spectra from the BALF, as well as hydrophilic and hydrophobic lung metabolites, enabled the high‐dose group to be discriminated from the control group; increased levels of isopropanol, ethane, and acetone and lower levels of ethanol, acetate, formate, and glycerophosphocholine were detected in the BALF of mice treated with higher doses of naphthalene. Furthermore, increased isopropanol and phosphorylcholine‐containing lipid levels and decreased succinate and glutamine levels were discovered in the lungs of naphthalene‐exposed mice. These metabolic changes may be related to lipid peroxidation, disruptions of membrane components and imbalanced energy supply, and these results may partially explain the loss of cell membrane integrity in the airway epithelial cells of naphthalene‐treated mice. We conclude that NMR‐based metabolomic studies on BALF and lung tissues are a powerful tool to understand the mechanisms underlying respiratory toxicity. Copyright © 2013 John Wiley & Sons, Ltd.
      PubDate: 2014-01-30T03:16:29.770367-05:
      DOI: 10.1002/jat.2970
       
  • Organ‐specific distribution of gold nanoparticles by their surface
           functionalization
    • Abstract: The behavior and fate of intravenously (i.v.) injected nanoparticles (NPs) can be controlled by several physicochemical factors including size, shape and surface charge. To evaluate the role of surface charge on distribution of NPs, we used neutral‐charged 15‐nm‐sized polyethylene glycol‐coated gold nanoparticles (AuNPPEG) as a core NP and carboxyl or amine groups were conjugated to AuNPPEG to generate negative (AuNPCOOH) or positive AuNP (AuNPNH2), respectively. Each type of AuNP was i.v. injected into mice (1 mg kg–1) and the concentration of Au was measured in different organs at 30 min, 4, 24 h, 7, 14 days, 1, 3 and 6 months post‐injection. The organ distribution also showed the higher deposition rate depending on their functional groups: AuNPPEG for mesenteric lymph node, kidney, brain and testis; AuNPCOOH for liver; AuNPNH2 for spleen, lung and heart. The blood circulation time and the major excretion route were different depending on their functional groups. In conclusion, functional groups conjugated on the surface of AuNPs produce differences in blood kinetics, organ distribution and elimination pattern which can be important information for directing NPs to specific organs or improving the kinetic properties. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Impacts of the feed contaminant deoxynivalenol on the intestine of
           monogastric animals: poultry and swine
    • Abstract: Deoxynivalenol (DON) is one of the most prevalent cereal contaminants with major public health concerns owing to its high toxigenic potentials. Once ingested, DON first and foremost targets epithelial cells of the gastrointestinal tract, whose proper functioning, as the first line of defence, is of paramount importance for the host's health. Emerging evidences, summarized in this article, suggest that DON produces its toxicity primarily via activation of the mitogen‐activated protein kinases (MAPKs) signalling pathway and alteration in the expression of genes responsible for key physiological and immunological functions of the intestinal tissue of chickens and pigs. The activation of MAPKs signalling cascade results in disruption of the gut barrier function and an increase in the permeability by reducing expression of the tight junction proteins. Exposure to DON also down‐regulates the expression of multiple transporter systems in the enterocytes with subsequent impairment of the absorption of key nutrients. Other major intestinal cytotoxic effects of DON described herein are modulation of mucosal immune responses, leading to immunosupression or stimulation of local immune cells and cytokine release, and also facilitation of the persistence of intestinal pathogens in the gut. Both of the last events potentiate enteric infections and local inflammation in pigs and poultry, rendering enterocytes and the host more vulnerable to luminal toxic compounds. This review highlights the cytotoxic risks associated with the intake of even low levels of DON and also identifies gaps of knowledge that need to be addressed by future research. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Cellular uptake and toxicity effects of silver nanoparticles in mammalian
           kidney cells
    • Abstract: The rapid progress and early commercial acceptance of silver‐based nanomaterials is owed to their biocidal activity. Besides embracing the antimicrobial potential of silver nanoparticles (AgNPs), it is imperative to give special attention to the potential adverse health effects of nanoparticles owing to prolonged exposure. Here, we report a detailed study on the in vitro interactions of citrate‐coated AgNPs with porcine kidney (Pk15) cells. As uncertainty remains whether biological/cellular responses to AgNPs are solely as a result of the release of silver ions or whether the AgNPs themselves have toxic effects, we investigated the effects of Ag+ on Pk15 cells for comparison. Next, we investigated the cellular uptake of both AgNPs and Ag+ in Pk15 cells at various concentrations applied. The detected Ag contents in cells exposed to 50 mg l−1 AgNPs and 50 mg l−1 Ag+ were 209 and 25 µg of Ag per 106 cells, respectively. Transmission electron microscopy (TEM) images indicated that the Pk15 cells internalized AgNPs by endocytosis. Both forms of silver, nano and ionic, decreased the number of viable Pk15 cells after 24 h in a dose‐dependent manner. In spite of a significant uptake into the cells, AgNPs had only insignificant toxicity at concentrations lower than 25 mg l−1, whereas Ag+ exhibited a significant decrease in cell viability at one‐fifth of this concentration. The Comet assay suggested that a rather high concentration of AgNP (above 25 mg l−1) is able to induce genotoxicity in Pk15 cells. Further studies must seek deeper understanding of AgNP behavior in biological media and their interactions with cellular membranes. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • d‐α‐tocopheryl polyethylene glycol 1000
           succinate‐containing vehicles provide no detectable chemoprotection
           from oxidative damage
    • Abstract: The objective of this study was to evaluate potential protective effects of vehicles containing d‐α‐tocopheryl polyethylene glycol 1000 succinate (TPGS), which may impact nonclinical safety assessments of oxidative processes. This was achieved by evaluating plasma, liver and adrenal gland concentrations of d‐α‐tocopheryl succinate (TS) and d‐α‐tocopherol as well as oxidative status of plasma following oral dosing of TPGS‐containing vehicles, intraperitoneal (IP) dosing of TS or ex vivo treatment of blood with H2O2. Male and female rats were dosed orally with formulations containing 5% or 40% TPGS (70 or 550 mg kg–1 day–1 TS, respectively) for 1 week. A control group was dosed orally with polyethylene glycol‐400 (PEG‐400; no vitamin E) and positive control animals received a single 100 mg kg–1 day–1 IP injection of TS. Whole blood from untreated animals was treated ex vivo with 5 or 50 mm H2O2, with or without TS (0.5, 5, 50 or 500 μm) or ascorbate (1 mm), for 1 h. Oral TPGS treatments did not affect d‐α‐tocopherol concentrations in plasma or adrenal glands and caused only transient increases in liver. Concentrations of TS in plasma, liver and adrenal glands were undetectable in control animals, but increased in all other groups. Oral administration of TPGS did not reduce plasma lipid peroxidation in vivo. Substantially greater TS concentrations used ex vivo (100× greater than in vivo) were also unable to reduce lipid peroxidation in H2O2‐treated whole blood. These results provide evidence that administration of oral TPGS vehicles is unlikely to impact nonclinical safety assessments of pharmaceuticals. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Comparison of the kinetics of various biomarkers of benzo[a]pyrene
           exposure following different routes of entry in rats
    • Abstract: The effect of route of exposure on the kinetics of key biomarkers of exposure to benzo[a]pyrene (BaP), a known human carcinogen, was studied. Rats were exposed to an intravenous, intratracheal, oral and cutaneous dose of 40 µmol kg–1 BaP. BaP and several metabolites were measured in blood, urine and feces collected at frequent intervals over 72 h post‐treatment, using high‐performance liquid chromatography/fluorescence. Only BaP and 3‐hydroxyBaP (3‐OHBaP) were detectable in blood at all time points. There were route‐to‐route differences in the excreted amounts (% dose) of metabolites but the observed time courses of the excretion rate were quite similar. In urine, total amounts of BaP metabolites excreted over the 0–72 h period followed the order: trans‐4,5‐dihydrodiolBaP (4,5‐diolBaP) ≥ 3‐OHBaP > 7‐OHBaP ≥ 7,8‐diolBaP after intravenous injection and intratracheal instillation; 3‐OHBaP ≈ 7‐OHBaP ≥ 4,5‐diolBaP > 7,8‐diolBaP after cutaneous application; 3‐OHBaP ≥ 4,5‐diolBaP ≈ 7‐OHBaP > 7,8‐diolBaP following oral administration. In feces, total amounts of BaP metabolites recovered were: 7‐OHBaP ≈ 3‐OHBaP > 4,5‐diolBaP > 7,8‐diolBaP > BaP‐7,8,9,10‐tetrol following all administration routes. For all exposure routes, excretion of 4,5‐ and 7,8‐diolBaP was almost complete over the 0–24 h period in contrast with that of 3‐ and 7‐OHBaP. This study confirms the interest of measuring multiple metabolites due to route‐to‐route differences in the relative excretion of the different biomarkers and in the time courses of diolBaPs versus OHBaPs. Concentration ratios of the different metabolites may help indicate time and main route of exposure. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Tween‐80 and impurity induce anaphylactoid reaction in zebrafish
    • Abstract: A number of recent reports suspected that Tween‐80 in injectable medicines, including traditional Chinese medicine injections could cause life‐threatening anaphylactoid reaction, but no sound conclusion was drawn. A drug‐induced anaphylactoid reaction is hard to be assayed in vitro and in conventional animal models. In this study, we developed a microplate‐based quantitative in vivo zebrafish assay for assessing anaphylactoid reaction and live whole zebrafish mast cell tryptase activity was quantitatively measured at a wavelength of 405 nm using N‐benzoyl‐dl‐arginine p‐nitroanilide as a substrate. We assessed 10 batches of Tween‐80 solutions from various national and international suppliers and three Tween‐80 impurities (ethylene glycol, 2‐chloroethanol and hydrogen peroxide) in this model and found that three batches of Tween‐80 (nos 2, 20080709 and 20080616) and one Tween‐80 impurity, hydrogen peroxide (H2O2), induced anaphylactoid reactions in zebrafish. Furthermore, we found that H2O2 residue and peroxide value were much higher in Tween‐80 samples 2, 20080709 and 20080616. These findings suggest that H2O2 residue in combination with oxidized fatty acid residues (measured as peroxide value) or more likely the oxidized fatty acid residues in Tween‐80 samples, but not Tween‐80 itself, may induce anaphylactoid reaction. High‐throughput zebrafish tryptase assay developed in this report could be used for assessing safety of Tween‐80‐containing injectable medicines and potentially for screening novel mast cell‐modulating drugs. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Animal models for percutaneous absorption
    • Abstract: Animal models are important tools to predict human in vivo percutaneous absorption/penetration. Monkey, pig, rat, rabbit, guinea pig, hairless rodents, such as hairless rat, hairless mouse, hairless guinea pig and hairless dog, are among the most used animals for this purpose. Each animal model has its own advantages and weakness or limitation. To better correlate animal data with human skin absorption, we need to be familiar with each animal model's characteristics as well as experimental method and condition. We reviewed the original papers published after 1993 that described permeability of both animal skin and human skin. It showed that monkey, pig and hairless guinea pig are more predictive of human skin absorption/penetration and common laboratory animals, such as rat, rabbit, guinea pig, generally overestimate human skin absorption/penetration. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Long‐term exposures to di‐n‐butyl phthalate inhibit body
           growth and impair gonad development in juvenile Murray rainbowfish
           (Melanotaenia fluviatilis)
    • Abstract: The aim of the present study was to evaluate whether long‐term exposures to environmentally relevant concentrations of di‐n‐butyl phthalate (DnBP) disrupt the reproduction‐based endpoints in juvenile Murray rainbowfish (Melanotaenia fluviatilis). Fish were exposed to 5, 15 or 50 µg l−1 DnBP for 30, 60 and 90 days each, and the effects on survival, body growth, whole‐body concentrations of sex steroid hormones and gonadal development were investigated. The lowest observed effective concentration to affect the condition factor after 90 days was 5 µg l−1. Complete feminization of the gonad was noted in fish exposed to 5 µg l−1 for 90 days and to 15 and 50 µg l−1 of DnBP for 30 or 60 days. After 90 days of exposure to DnBP, the ovaries were regressed and immature as opposed to the control fish which were in early‐vitellogenic stage. Testes, present only in fish exposed to 5 µg l−1 of DnBP for 30 or 60 days, were immature in comparison to the control fish that contained testes in the mid‐spermatogenic phase. The E2/11‐KT ratio was significantly higher only after exposures to 5 µg l−1 DnBP for 90 days and 50 µg l−1 DnBP for 30 days. Our data suggest that exposures to 5 µg l−1 DnBP for 30 days did not have profound effects on body growth and gonadal differentiation of fish. However, 30 days of exposure to 15 µg l−1 could interfere with the gonad development and to 50 µg l−1 could compromise the hormonal profile of juvenile fish. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Impact of di‐ethylhexylphthalate exposure on metabolic programming
           in P19 ECC‐derived cardiomyocytes
    • Abstract: Di(2‐ethylhexyl)phthalate (DEHP) is the most common plasticizer in plastic devices of everyday use. It is a ubiquitous environmental contaminant and primarily known to impair male gonadal development and fertility. Studies concerning the long‐term effects of prenatal DEHP exposure on certain diseases [The Developmental Origins of Health and Disease paradigm (DOHaD) hypothesis] are scarce although it is proven that DEHP crosses the placenta. Rising environmental pollution during the last centuries coincides with an increasing prevalence of cardiovascular and metabolic diseases. We have investigated the effects of an early embryonic DEHP exposure at different developmental stages on cardiomyogenesis. We used an in‐vitro model, the murine P19 embryonic carcinoma cell line (P19 ECC), mimicking early embryonic stages up to differentiated beating cardiomyocytes. P19 ECC were exposed to DEHP (5, 50, 100 µg ml–1) at the undifferentiated stage for 5 days and subsequently differentiated to beating cardiomyocytes. We analyzed the expression of metabolic (Pparg1, Fabp4 and Glut4), cardiac (Myh6, Gja1) and methylation (Dnmt1, Dnmt3a) marker genes by quantitative real‐time PCR (qRT‐PCR), beating rate and the differentiation velocity of the cells. The methylation status of Pparg1, Ppara and Glut4 was investigated by pyrosequencing. DEHP significantly altered the expression of all investigated genes. The beating rate and differentiation velocity were accelerated. Exposure to DEHP led to small but statistically significant increases in methylation of specific CpGs within Ppara and Pparg1, which otherwise were generally hypomethylated, but methylation of Glut4 was unaltered. Early DEHP exposure of P19 ECC alters the expression of genes associated with cellular metabolism and the functional features of cardiomyocytes. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Assessment of temperature‐induced hERG channel blockade variation by
           drugs
    • Abstract: Drug‐induced QT prolongation has been reported in humans and animals. This potentially lethal effect can be induced by drugs interacting with a cardiac potassium channel, namely hERG (human ether‐a go‐go‐related gene) leading to arrhythmia or torsade de pointes (TdP). Hence, in vitro evaluation of therapeutics for their effects on the rapid delayed rectifier current (IKr) mediated by the K+ ion channel encoded by hERG is a valuable tool for identifying potential arrhythmic side effects during drug safety testing. Our objective was to evaluate the temperature‐induced hERG channel blockade variation by human and veterinary drugs using the IonFlux 16 system. A panel of eight drugs was tested for IKr inhibition at both ambient (23 °C) and physiological (37 °C) temperatures at various concentrations using IonFlux 16, an automated patch clamp system. Our results established that both amiodarone (IC50 = 0.56 μM at 23 °C and 0.30 μM at 37 °C) and β‐estradiol (IC50 = 24.72 μM at 23 °C and 8.17 μM at 37 °C) showed a dose‐dependent IKr blockade with a higher blockade at 37 °C. Whereas, blockade of IKr by both ivermectin (IC50 = 12.52 μM at 23 °C and 24.41 μM at 37 °C) and frusemide (IC50 = 12.58 μM at 23 °C and 25.55 μM at 37 °C) showed a dose‐dependent IKr blockade with a lower blockade at 37 °C. Gentamicin, enrofloxacin, xylazine and albendazole did not block IKr at both the assessed temperatures. Collectively, these results demonstrate that the effect of temperature variation should be taken into consideration during the evaluation of test drugs for their hERG channel blockade potential. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Use of the mouse ear vesicant model to evaluate the effectiveness of
           ebselen as a countermeasure to the nitrogen mustard mechlorethamine
    • Abstract: Previous studies in this and other laboratories have demonstrated that ebselen (EB‐1), an organoselenium compound, spares cells from mechlorethamine (HN2) toxicity in vitro. In the present study, the hypothesis that EB‐1 will reduce dermal toxicity of HN2 in vivo is put forward and found to have merit. Using the mouse ear vesicant model (MEVM), HN2, applied topically, showed a dose‐dependent effect upon ear swelling and thickness 24 h after treatment; whereas tissue injury consistent with vesication was observed at the higher test doses of HN2 (≥ 0.250 µmol per ear). To examine HN2 countermeasure activity using the MEVM, either hydrocortisone (HC), as a positive control, or EB‐1, the test countermeasure, was administered as three topical treatments 15 min, 4 and 8 h after HN2 exposure. Using this approach, both HC and EB‐1 were found to reduce tissue swelling associated with HN2 toxicity 24 h after exposure to the vesicant. Taken together, these data demonstrate for the first time the effectiveness of EB‐1 as a vesicant countermeasure in a relevant in vivo model. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Gene expression profile of brain regions reflecting aberrations in nervous
           system development targeting the process of neurite extension of rat
           offspring exposed developmentally to glycidol
    • Abstract: We previously found that exposure to glycidol at 1000 ppm in drinking water caused axonopathy in maternal rats and aberrations in late‐stage hippocampal neurogenesis, targeting the process of neurite extension in offspring. To identify the profile of developmental neurotoxicity of glycidol, pregnant Sprague–Dawley rats were given drinking water containing glycidol from gestational day 6 until weaning on day 21 after delivery, and offspring at 0, 300 and 1000 ppm were subjected to region‐specific global gene expression profiling. Four brain regions were selected to represent both cerebral and cerebellar tissues, i.e., the cingulate cortex, corpus callosum, hippocampal dentate gyrus and cerebellar vermis. Downregulated genes in the dentate gyrus were related to axonogenesis (Nfasc), myelination (Mal, Mrf and Ugt8), and cell proliferation (Aurkb and Ndc80) at ≥ 300 ppm, and upregulated genes were related to neural development (Frzb and Fzd6) at 1000 ppm. Upregulation was observed for genes related to myelination (Kl, Igf2 and Igfbp2) in the corpus callosum and axonogenesis and neuritogenesis (Efnb3, Tnc and Cd44) in the cingulate cortex, whereas downregulation was observed for genes related to synaptic transmission (Thbs2 and Ccl2) in the cerebellar vermis; all of these changes were mostly observed at 1000 ppm. Altered gene expression of Cntn3, which functions on neurite outgrowth‐promotion, was observed in all four brain regions at 1000 ppm. Gene expression profiles suggest that developmental exposure to glycidol affected plasticity of neuronal networks in the broad brain areas, and dentate gyrus neurogenesis may be the sensitive target of this type of toxicity. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • A longitudinal study assessing lens thickness changes in the eye of the
           growing beagle using ultrasound scanning: relevance to age of dogs in
           regulatory toxicology studies
    • Abstract: The lens is formed in utero with new secondary lens fibres added as outer layers throughout life in a growth pattern characteristic of the species. This study examined the time course of beagle lens growth to better understand the optimal starting age of dogs for safety studies to support adult versus paediatric indications, and to assess the feasibility of non‐invasively monitoring lens growth with high frequency ultrasound. Ultrasound scanning was performed in six female beagle dogs using the Vevo770. All dogs were imaged in B‐mode using local anaesthetic but without sedation. Imaging was carried out every 2 weeks from 8 to 22 weeks of age and then monthly until 62 weeks of age. The dogs tolerated the procedure well. The lens was visible in all dogs and measuring the lens thickness with high frequency ultrasound demonstrated good analytical reproducibility [Root Mean Square (RMS) = 3.13%]. No differences between the left and right eye existed and lens thickness correlated with body weight. The highest weekly growth rate was before 12 weeks of age. A statistically significant difference between monthly thickness was detected until 42 weeks of age at which point growth reached a plateau. During the experiment, lenses grew by 29.7% reaching an average thickness of 6.4 mm ± 0.03. By 10 months of age (the typical age used for routine toxicological evaluation), beagles have reached a plateau in lens growth that is analogous to human adults. Where lens is a target organ of concern it is suggested that beagles under 6 months old may be a better model for determining paediatric safety. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Absence of mature microRNAs inactivates the response of gene expression to
           carcinogenesis induced by N‐ethyl‐N‐nitrosourea in mouse
           liver
    • Abstract: This study aims to evaluate the role of microRNAs (miRNAs) in chemical tumorigenesis by evaluating genomic gene expression in miRNA knockout mice. Previous studies showed that mice without mature miRNAs due to hepatocyte‐specific Dicer1 knockout (KO) had a much higher liver tumor incidence than wild‐type mice. In this study, Dicer1 KO or the wild‐type mice were treated intraperitoneally with genotoxic carcinogen N‐ethyl‐N‐nitrosourea (ENU) at a single dose (150 mg kg–1 that resulted in liver tumorigenesis) or the vehicle at 3 weeks of age. The animals were killed 2 weeks after treatment and the liver samples were collected for the gene expression study. Principal components analysis and hierarchical cluster analysis showed that gene expression was globally altered by the Dicer1 KO and ENU exposure. There were 5621, 3286 and 2565 differentially expressed genes for Dicer1 disruption, ENU treatment in wild‐type mice and ENU treatment in Dicer1 KO mice, respectively. Functional analysis of the differentially expressed genes suggests that the Dicer1 KO mouse liver lost their capability to suppress the carcinogenesis induced by ENU exposure in genomic level. In addition, the miRNA‐mediated BRCA1 and P53 signaling pathways were identified as the main pathways responsible for the tumorigenesis. We conclude that the mouse livers in the absence of mature miRNAs could not appropriately respond to carcinogenic insults from ENU treatment, indicating that miRNAs play a critical role in chemical carcinogenesis. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Application of the SOS/umu test and high‐content in vitro
           micronucleus test to determine genotoxicity and cytotoxicity of nine
           benzothiazoles
    • Abstract: Benzothiazole and benzothiazole derivatives (BTs) have been detected in various environmental matrices as well as in human beings, but little is currently available regarding their toxicities. In our study, genotoxicities of nine BTs (benzothiazole [BT], 2‐chlorobenzothiazole [CBT], 2‐bromobenzothiazole [BrBT], 2‐fluorobenzothiazole [FBT], 2‐methylbenzothiazole [MeBT], 2‐mercaptobenzothiazole [MBT], 2‐aminobenzothiazole [ABT], 2‐hydroxy‐benzothiazole [OHBT] and 2‐methythiobenzothiazole [MTBT]) are comprehensively evaluated by the SOS/umu test using the bacterial Salmonella typhimurium TA1535/pSK1002 for DNA‐damaging effect and the high content in vitro micronucleus test using two human carcinoma cells (MGC‐803 and A549) for chromosome‐damaging effect. The cytotoxicity of BTs on both bacteria and two human cells was also evaluated. Except for the cytotoxic effect of MBT on MGC‐803 and A549, the other tested BTs showed more than 50% cytotoxicity at their highest concentrations in a dose‐dependent manner, and their LC50s ranged from 19 (MBT in bacteria) to 270 mg l–1 (CBT in A549). Activation and inactivation were observed for specific BTs after metabolism. On the other hand, no evidence of genotoxicity was obtained for BT, FBT and MBT, and DNA damage was induced by ABT, OHBT, BrBT and MTBT in MGC‐803, by MeBT in A549 and by CBT in both cells. Through quantitative structure–activity relationship analysis, two structure alerts for chemical genotoxicity, including heterocyclic amine and hacceptor‐path3‐hacceptor are present in ABT and OHBT respectively; however, the underlying mechanisms still need further evaluation. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • CKD‐501, a novel selective PPARγ agonist, shows no carcinogenic
           potential in ICR mice following oral administration for 104 weeks
    • Abstract: CKD‐501 is a peroxisome proliferator‐activated receptor gamma (PPARγ) agonist that is effective for the treatment of diabetes. However, its carcinogenic potential remains controversial. The current carcinogenicity study was conducted over a period of 104 weeks in ICR mice. Three groups, each consisting of 60 male and 60 female mice, received oral CKD‐501 dosages of 0.2, 1.0 or 6.0 mg kg−1day–1. The mortality rates of the male control, 0.2, 1.0 and 6.0 mg kg–1 day–1 treated groups were 60%, 68%, 58% and 67%, respectively and 57%, 68% and 67% in the female control, 0.2 and 1.0 mg kg−1 day–1 treated groups. It was 67% in the female 6.0 mg kg−1 day–1 treated group, which was terminated at week 98 due to its increased mortality rate. No significant treatment‐related effects were observed on the survival rates, with the exception of females in the 6.0 mg kg−1 day–1 group. Body weights increased in females receiving 1.0 and 6.0 mg kg−1 day–1 due to the class effects of the PPARγ agonist. Differences were not found in hematology parameters between the CKD‐501‐treated groups and their corresponding controls, but the histopathological evidence did not reveal any findings attributed to CKD‐501. Treated animals exhibited non‐neoplastic findings (adipocyte proliferation, bone marrow hypoplasia cardiomyopathy), but all of these were expected changes for this class of compound. There were no treatment‐related neoplastic changes in this study. The results of this study therefore demonstrate a lack of carcinogenicity following oral administration of CKD‐501 to ICR mice for 104 weeks. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • The immunosuppressant tributyltin oxide blocks the mTOR pathway, like
           rapamycin, albeit by a different mechanism
    • Abstract: We treated the thymoma cell line (EL4) with two model immunosuppressants, rapamycin and tributyltin oxide (TBTO), and compared their effects on the expression levels of proteins that are downstream targets of mTOR kinase 1 (mammalian target of rapamycin, known also as mechanistic target of rapamycin): p70 ribosomal S6 kinase1 and 4E‐binding protein 1, a repressor of the cap‐binding protein eIF4E. In addition, we evaluated the levels of ribosomal protein S6, p‐eIF4B, substrates of p70S6 kinase1, matrin 3 and ribonucleotide reductase, subunit RRM2. The levels of these proteins were evaluated in cell lysates by immunoblot. We found that both compounds inhibited the phosphorylation state of p70S6 kinase 1 and its substrates; however, TBTO, in contrast to rapamycin, reduced the level of the total p70S6k1. Besides, we detected a band with a molecular weight of c. 32 kDa only in the TBTO‐treated lysates. This band was detected with a monoclonal antibody specific for S6k1, suggesting that this band might be a degradation product of the kinase. Further, TBTO and rapamycin differentially affected 4E‐binding protein 1; the former compound stimulated its phosphorylation state whereas the latter inhibited it. The two immunosuppressants did not affect the level of ribonucleotide reductase, but TBTO downregulated matrin3, in agreement with a previous report, whereas rapamycin had no effect on the expression level of this latter protein. We conclude that TBTO inhibits, like rapamycin, the p70 S6 kinase 1 pathway, but with a different mechanism. However, in contrast to rapamycin, which inhibits the cap‐dependent translation, TBTO increases the phosphorylation of 4E‐binding protein1. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Proteomic analysis of perfluorooctane sulfonate‐induced apoptosis in
           human hepatic cells using the iTRAQ technique
    • Abstract: Perfluorooctane sulfonate (PFOS) is one of the most commonly used perfluorinated compounds, whose environmental exposure has been associated with a number of adverse health outcomes. However, the molecular mechanisms involved in PFOS toxicity are still not well elucidated. In the present study, we applied iTRAQ labeling quantitative proteomic technology to investigate the differential protein expression profiles of non‐tumor human hepatic cells (L‐02) exposed to PFOS. A total of 18 proteins were differentially expressed in a dose‐dependent manner in PFOS‐treated cells versus the control. Among these, 11 proteins were up‐regulated and 7 were down‐regulated. Gene ontology analysis indicated that PFOS would exert toxic effects on L‐02 cells by affecting multiple biological processes, including protein biosynthesis and degradation, mRNA processing and splicing, transcription, signal transduction and transport. Furthermore, the proteomic results especially proposed that the inhibition of HNRNPC, HUWE1 and UBQLN1, as well as the induction of PAF1 is involved in the activation of the p53 and c‐myc signaling pathways, which then trigger the apoptotic process in L‐02 cells exposed to PFOS. Overall, these data will aid our understanding of the mechanisms responsible for PFOS‐mediated hepatotoxicity, and develop useful biomarkers for monitoring and evaluating PFOS contamination in the environment. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Identification of differentially expressed proteins related to
           organophosphorus‐induced delayed neuropathy in the brains of hens
    • Abstract: Some organophosphorus compounds can cause organophosphate‐induced delayed neuropathy (OPIDN). Incidents have been documented for decades, however, little is known about which proteins contribute to the initiation, progression and development of OPIDN. In this study, 51 hens were divided into three groups. The tri‐ortho‐cresyl‐phosphate (TOCP) group was treated with 1000 mg kg–1 TOCP whereas the control group was treated with an equivalent volume of vehicle. The PMSF + TOCP group was treated subcutaneously with 40 mg kg–1 phenylmethylsulfonyl fluoride (PMSF), followed by 1000 mg kg–1 TOCP 24 h later. Proteins in the brains of hens were separated by two‐dimensional polyacrylamide gel electrophoresis on day 5 after TOCP administration. Mass spectrometry identified eight differentially expressed proteins. Among these proteins, downregulated expression of glutamine synthetase (GS) in the brains of hens after TOCP treatment was further confirmed by real time RT‐PCR and ELISA. Moreover, the brains of hens exposed to TOCP exhibited increased levels of glutamate (Glu) and cytosolic calcium concentration ([Ca2+]i), and a decreased level of glutamine (Gln). However, there were no significant differences in GS expression or levels of Glu, Gln, and [Ca2+]i in the brains of hens among the groups on day 21 after TOCP administration. These results indicate that TOCP exposure downregulates GS expression in the brains of hens, and that downregulation of GS is accompanied by increased levels of Glu and [Ca2+]i in the early stage after TOCP administration. It is also suggested that the downregulated expression of GS might be associated with OPIDN through the disruption of homeostasis of the Glu–Gln cycle and [Ca2+]i. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Combined antitumor effects of bee venom and cisplatin on human cervical
           and laryngeal carcinoma cells and their drug resistant sublines
    • Abstract: In the present study, we investigated the possible combined anticancer ability of bee venom (BV) and cisplatin towards two pairs of tumour cell lines: parental cervical carcinoma HeLa cells and their cisplatin‐resistant HeLa CK subline, as well as laryngeal carcinoma HEp‐2 cells and their cisplatin‐resistant CK2 subline. Additionally, we identified several peptides of BV in the BV sample used in the course of the study and determined the exact concentration of MEL. BV applied alone in concentrations of 30 to 60 µg ml–1 displayed dose‐dependent cytotoxicity against all cell lines tested. Cisplatin‐resistant cervical carcinoma cells were more sensitive to BV than their parental cell lines (IC50 values were 52.50 µg ml–1 for HeLa vs. 47.64 µg ml–1 for HeLa CK cells), whereas opposite results were obtained for cisplatin‐resistant laryngeal carcinoma cells (IC50 values were 51.98 µg ml–1 for HEp‐2 vs. > 60.00 µg ml–1 for CK2 cells). Treatment with BV alone induced a necrotic type of cell death, as shown by characteristic morphological features, fast staining with ethidium‐bromide and a lack of cleavage of apoptotic marker poly (ADP‐ribose) polymerase (PARP) on Western blot. Combined treatment of BV and cisplatin induced an additive and/or weak synergistic effect towards tested cell lines, suggesting that BV could enhance the killing effect of selected cells when combined with cisplatin. Therefore, a greater anticancer effect could be triggered if BV was used in the course of chemotherapy. Our results suggest that combined treatment with BV could be useful from the point of minimizing the cisplatin concentration during chemotherapy, consequently reducing and/or postponing the development of cisplatin resistance. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Protective effects of schizandrin and schizandrin B towards cisplatin
           nephrotoxicity in vitro
    • Abstract: Renal proximal tubular epithelial cells are the main targets of toxic drugs such as cisplatin (CisPt), an alkylating agent indicated for the treatment of solid organ tumors. Current techniques aiming at reducing nephrotoxicity in patients receiving CisPt are still not satisfactory as they can only partially prevent acute kidney injury. New nephroprotective strategies remain to be developed. In the present in vitro study, schizandrin (Schi) and schizandrin B (Schi B), major phytochemicals from Schisandra chinensis (Turcz.) Baill. fruits, were tested on HK‐2 cells along four processes that could help alleviate CisPt toxicity. Results indicated that: (i) both Schi and Schi B enhanced cell survival via reducing apoptosis rate; (ii) only Schi showed moderate effects towards modulation of regeneration capacities of healthy cells; (iii) both Schi and Schi B limited extracellular matrix deposition; and (iv) both compounds could help preventing dedifferentiation processes via the β‐catenin pathway. Schi and Schi B present promising activities for future development of protective agents against CisPt nephrotoxicity. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Hepatic fibrogenesis and transforming growth factor/Smad signaling
           activation in rats chronically exposed to low doses of lead
    • Abstract: Lead is an important heavy metal pollutant in the environment. The nervous system, kidney and liver are the most susceptible organs to lead deposition, showing that this pollutant has no single target system. To examine the cellular and molecular mechanisms involved in their pathobiology of chronic lead at low‐dose exposure in the liver, male Wistar rats were exposed to 0.06% lead acetate in drinking water every day for 4 months. At the end of the study, hepatic metal accumulation, morphology and function were examined. Immunochemical staining and Western blot analysis were performed to detect extracellular matrix proteins, α‐smooth muscle actin and transforming growth factor (TGF)β1/Smad pathway expression. Results showed increased laminin, collagen IV and fibronectin, located at the perisinusoidal space. Phenotypic transformation of hepatic stellate cells into myofibroblast‐like cells was evidenced at the ultrastructural level and a significant expression of α‐smooth muscle actin in Disse's space was observed. These findings were associated with a marked increase in TGFβ1/Smad2/3 signaling. Our data suggest that, chronically, exposure to low levels of lead could trigger the onset of a hepatic fibrogenic process through upregulated TGFβ1/Smad signaling. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Histopathological findings on Carassius auratus hepatopancreas upon
           exposure to acrylamide: correlation with genotoxicity and metabolic
           alterations
    • Abstract: Acrylamide is an amide used in several industrial applications making it easily discharged to aquatic ecosystems. The toxicity of acrylamide to aquatic organisms is scarcely known, although previous studies with murine models provided evidence for deleterious effects. To assess the effects of acrylamide to freshwater fish, goldfish (Carassius auratus L.) were exposed to several concentrations of waterborne acrylamide and analysed for genotoxic damage, alterations to detoxifying enzymes and histopathology. Results revealed a dose‐dependent increase in total DNA strand breakage, the formation of erythrocytic nuclear abnormalities and in the levels of hepatic cytochrome P4501A (CYP1A) and glutathione S‐transferase (GST) activity. In addition, acrylamide induced more histopathological changes to pancreatic acini than to the hepatic parenchyma, regardless of exposure concentration, whereas hepatic tissue only endured significant alterations at higher concentrations of exposure. Thus, results confirm the genotoxic potential of acrylamide to fish and its ability to induce CYP1A, probably as a direct primary defence mechanism. This strongly suggests the substance's pro‐mutagenic potential in fish, similarly to what is known for rodents. However, the deleterious effects observed in the pancreatic acini, more severe than in the liver, could indicate a specific, albeit unknown toxic mechanism of acrylamide to fish that overran the organism's metabolic defences against a chemical agent rather than causing a general systemic failure. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Alteration in cellular viability, pro‐inflammatory cytokines and
           nitric oxide production in nephrotoxicity generation by Amphotericin B:
           involvement of PKA pathway signaling
    • Abstract: Amphotericin B is one of the most effective antifungal agents; however, its use is often limited owing to adverse effects, especially nephrotoxicity. The purpose of this study was to evaluate the effect of inhibiting the PKA signaling pathway in nephrotoxicity using Amphotericin B from the assessment of cell viability, pro‐inflammatory cytokines and nitric oxide (NO) production in LLC‐PK1 and MDCK cell lines. Amphotericin B proved to be cytotoxic for both cell lines, as assessed by the mitochondrial enzyme activity (MTT) assay; caused DNA fragmentation, determined by flow cytometry using the propidium iodide (PI) dye; and activated the PKA pathway (western blot assay). In MDCK cells, the inhibition of the PKA signaling pathway (using the H89 inhibitor) caused a significant reduction in DNA fragmentation. In both cells lines the production of interleukin‐6 (IL)‐6 proved to be a dependent PKA pathway, whereas tumor necrosis factor‐alpha (TNF‐α) was not influenced by the inhibition of the PKA pathway. The NO production was increased when cells were pre‐incubated with H89 followed by Amphotericin B, and this production produced a dependent PKA pathway in LLC‐PK1 and MDCK cells lines. Therefore, considering the present study's results as a whole, it can be concluded that the inhibition of the PKA signaling pathway can aid in reducing the degree of nephrotoxicity caused by Amphotericin B. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Deltamethrin induced an apoptogenic signalling pathway in murine
           thymocytes : exploring the molecular mechanism
    • Abstract: Deltamethrin (DLM) is a well‐known pyrethroid insecticide; however, the immunotoxic effects of DLM on the mammalian system and its mechanism is still unclear. This study has been designed to first observe the binding affinity of DLM to immune cell receptors and its effects on the immune system. The docking score revealed that DLM has a strong binding affinity towards the CD4 and CD8 receptors. DLM induces apoptosis in murine thymocytes in a concentration‐dependent manner. The ear\ly markers of apoptosis such as enhanced reactive oxygen species (ROS) and caspase‐3 activation are evident as early as 1 h by 25 and 50 μM DLM. Glutathione (GSH) depletion has also been observed at 1 h by 50 μM DLM concentration. In cell‐cycle studies using flow cytometry, the fraction of hypodiploid cells has gradually increased with all the concentrations of DLM at 18 h. The Annexin V binding assay measures the effect of DLM on apoptotic and necrotic cells. The apoptotic cells raised from 18.6% to 35.21% (10–50 μM DLM) at 18 h. N‐acetyl cysteine (NAC) effectively reduced the percentage of apoptotic cells which is increased by DLM. In contrast, buthionine sulfoximine (BSO) caused an elevation in the percentage of apoptotic cells. These results demonstrate that caspase activation, ROS activation and GSH act as critical mediators in a DLM‐induced apoptogenic signalling pathway in murine thymocytes. In the presence of caspase inhibitor, the percentage of apoptotic cells is partially decreased. Thus, there may be the possibility of some other caspase‐independent pathways in DLM‐induced apoptosis. Copyright © 2013 John Wiley & Sons, Ltd.
       
  • Evaluation of electric arc furnace‐processed steel slag for dermal
           corrosion, irritation, and sensitization from dermal contact
    • Abstract: Electric arc furnace (EAF) steel slag is alkaline (pH of ~11–12) and contains metals, most notably chromium and nickel, and thus has potential to cause dermal irritation and sensitization at sufficient dose. Dermal contact with EAF slag occurs in many occupational and environmental settings because it is used widely in construction and other industrial sectors for various applications including asphaltic paving, road bases, construction fill, and as feed for cement kilns construction. However, no published study has characterized the potential for dermal effects associated with EAF slag. To assess dermal irritation, corrosion and sensitizing potential of EAF slag, in vitro and in vivo dermal toxicity assays were conducted based on the Organisation for Economic Co‐operation and Development (OECD) guidelines. In vitro dermal corrosion and irritation testing (OECD 431 and 439) of EAF slag was conducted using the reconstructed human epidermal (RHE) tissue model. In vivo dermal toxicity and delayed contact sensitization testing (OECD 404 and 406) were conducted in rabbits and guinea pigs, respectively. EAF slag was not corrosive and not irritating in any tests. The results of the delayed contact dermal sensitization test indicate that EAF slag is not a dermal sensitizer. These findings are supported by the observation that metals in EAF slag occur as oxides of low solubility with leachates that are well below toxicity characteristic leaching procedure (TCLP) limits. Based on these results and in accordance to the OECD guidelines, EAF slag is not considered a dermal sensitizer, corrosive or irritant. Copyright © 2014 John Wiley & Sons, Ltd.
       
  • Further studies toward a mouse model for biochemical assessment of
           neuropathic potential of organophosphorus compounds
    • Abstract: Inhibition and aging of neuropathy target esterase (NTE) by neuropathic organophosphorus (OP) compounds triggers OP compound‐induced delayed neuropathy (OPIDN), whereas inhibition of acetylcholinesterase (AChE) produces cholinergic toxicity. The neuropathic potential of an OP compound is defined by its relative inhibitory potency toward NTE vs. AChE assessed by enzyme assays following dosing in vivo or after incubations of direct‐acting compounds or active metabolites with enzymes in vitro. The standard animal model of OPIDN is the adult hen, but its large size and high husbandry costs make this species a burdensome model for assessing neuropathic potential. Although the mouse does not readily exhibit clinical signs of OPIDN, it displays axonal lesions and expresses brain AChE and NTE. Therefore, the present research was performed as a further test of the hypothesis that inhibition of mouse brain AChE and NTE could be used to assess neuropathic potential using mouse brain preparations in vitro or employing mouse brain assays following dosing of OP compounds in vivo. Excellent correlations were obtained for inhibition kinetics in vitro of mouse brain enzymes vs. hen brain and human recombinant enzymes. Furthermore, inhibition of mouse brain AChE and NTE after dosing with OP compounds afforded ED50 ratios that agreed with relative inhibitory potencies assessed in vitro. Taken together, results with mouse brain enzymes demonstrated consistent correspondence between in vitro and in vivo predictors of neuropathic potential, thus adding to previous studies supporting the validity of a mouse model for biochemical assessment of the ability of OP compounds to produce OPIDN. Copyright © 2014 John Wiley & Sons, Ltd.
       
 
 
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