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  Subjects -> ENVIRONMENTAL STUDIES (Total: 786 journals)
    - ENVIRONMENTAL STUDIES (722 journals)
    - POLLUTION (21 journals)
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ENVIRONMENTAL STUDIES (722 journals)            First | 1 2 3 4 5 6 7 8     

Interdisciplinary Environmental Review     Hybrid Journal   (Followers: 4)
Interfaces     Full-text available via subscription   (Followers: 6)
International Aquatic Research     Open Access   (Followers: 3)
International Archives of Occupational and Environmental Health     Hybrid Journal   (Followers: 4)
International Environmental Agreements: Politics, Law and Economics     Hybrid Journal   (Followers: 11)
International Gambling Studies     Hybrid Journal   (Followers: 6)
International Innovation - climate     Open Access   (Followers: 1)
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: 2)
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: 9)
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: 2)
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: 6)
International Journal of Environment and Pollution     Hybrid Journal   (Followers: 5)
International Journal of Environment and Sustainable Development     Hybrid Journal   (Followers: 16)
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: 2)
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: 15)
International Journal of Environmental Science and Technology     Hybrid Journal   (Followers: 5)
International Journal of Environmental Studies     Hybrid Journal   (Followers: 10)
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: 5)
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: 2)
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: 6)
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: 9)
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: 7)
International Journal of Water Resources and Environmental Engineering     Open Access   (Followers: 1)
International Review of Environmental and Resource Economics     Full-text available via subscription   (Followers: 1)
International Studies in the Philosophy of Science     Hybrid Journal   (Followers: 9)
Interventions : International Journal of Postcolonial Studies     Hybrid Journal   (Followers: 9)
IOP Conference Series: Earth and Environmental Science     Open Access   (Followers: 7)
Iranian Studies     Hybrid Journal   (Followers: 9)
Irish Educational Studies     Hybrid Journal   (Followers: 2)
Irish Journal of Earth Sciences     Full-text available via subscription  
Irish Political Studies     Hybrid Journal   (Followers: 9)
ISLE: Interdisciplinary Studies in Literature and Environment     Hybrid Journal   (Followers: 1)
Isotopes in Environmental and Health Studies     Hybrid Journal   (Followers: 1)
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 Advanced Research in Civil and Environmental Engineering     Open Access   (Followers: 1)
Journal of Advances in Environmental Health Research     Open Access  
Journal of Agricultural and Environmental Ethics     Hybrid Journal   (Followers: 9)
Journal of Agricultural Biotechnology and Sustainable Development     Open Access  
Journal of Agricultural Chemistry and Environment     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: 251)
Journal of Applied Meteorology and Climatology     Full-text available via subscription   (Followers: 8)
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: 10)
Journal of Applied Volcanology     Open Access   (Followers: 8)
Journal of Arid Environments     Hybrid Journal   (Followers: 8)
Journal of Asian Studies     Full-text available via subscription   (Followers: 24)
Journal of Biochemical and Molecular Toxicology     Hybrid Journal   (Followers: 4)
Journal of Black Studies     Hybrid Journal   (Followers: 2)
Journal of Chemical Ecology     Hybrid Journal   (Followers: 2)

  First | 1 2 3 4 5 6 7 8     

Journal Cover   Journal of Applied Toxicology
  [SJR: 0.689]   [H-I: 47]   [12 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  [1606 journals]
  • Two‐generation reproduction and teratology studies of feeding
           aditoprim in Wistar rats
    • Authors: Xu Wang; Ziqiang Tan, Guyue Cheng, Ihsan Awais, Lingli Huang, Dongmei Chen, Yuanhu Pan, Zhenli Liu, Zonghui Yuan
      Pages: n/a - n/a
      Abstract: Aditoprim, a new bacteriostatic agent that belongs to diaminopyrimidines, has a broad antimicrobial spectrum, good antibacterial activity and excellent pharmacokinetics. To evaluate the reproductive toxicity and teratogenic potential of aditoprim, different concentrations of aditoprim were administered to Wistar rats by feeding diets containing 0, 20, 100 and 1000 mg kg–1, respectively. Each group consisting of 18 males and 25 females (F0) was treated with different concentrations of aditoprim through a 13‐week period before mating and during mating, gestation, parturition and lactation. At weaning, 20 males and 25 females of the F1 generation weanlings per group were selected randomly as parents for the F2 generation. Selected F1 weanlings were exposed to the same diet and treatment as their parents. At 1000 mg kg–1 dose group, body weights in F0 and F1 rats, fetal body weight on day 21 (0, 4 and 21) after birth and number of viable fetuses in the F0 and F1 generation significantly decreased. Teratogenicity study was performed in combination with the F1 generation of a two‐generation reproduction study. F1 parents of the reproduction study were mated after weaning of the F2a pups. Pregnant female rats were subjected to cesarean section on gestational day 20 for teratogenic examination. At 1000 mg kg–1 group, body weights, fetal body lengths, tail lengths, litter weights and number of viable fetuses were significantly decreased. No obvious external, skeletal or visceral malformations in fetuses were noted in any groups in the teratogenic test. The no‐observed‐adverse‐effect level for reproduction/development toxicity of aditoprim was 100 mg kg–1 diet (about 7.89–9.25 mg kg–1 body weight day–1). Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-07T02:26:45.347717-05:
      DOI: 10.1002/jat.3121
  • Human bone morphogenetic protein‐7 does not counteract aristolochic
           acid‐induced renal toxicity
    • Authors: Marie‐Hélène Antoine; Frédéric Debelle, Julie Piccirilli, Fadoua El Kaddouri, Anne‐Emilie Declèves, Eric De Prez, Cécile Husson, Frédérique Mies, Marie‐Françoise Bourgeade, Joëlle L. Nortier
      Pages: n/a - n/a
      Abstract: Aristolochic acids (AA) are nephrotoxic and profibrotic agents, leading to chronic kidney disease. As some controversial studies have reported a nephroprotective effect of exogenous recombinant human bone morphogenetic protein (rhBMP)‐7 in several models of renal fibrosis, we investigated the putative effect of rhBMP‐7 to prevent progressive tubulointerstitial damage after AA intoxication in vitro and in vivo. In vitro, the toxicity of AA on renal tubular cells was demonstrated by an increase in vimentin as well as a decrease in β‐catenin expressions, reflecting a dedifferentiation process. Increased fibronectin and interleukin‐6 levels were measured in the supernatants. Enhanced α‐SMA mRNA levels associated to decreased E‐cadherin mRNA levels were also measured. Incubation with rhBMP‐7 only prevented the increase in vimentin and the decrease in β‐catenin expressions. In vivo, in a rat model of AA nephropathy, severe tubulointerstitial lesions induced by AA after 10 and 35 days (collagen IV deposition and tubular atrophy), were not prevented by the rhBMP‐7 treatment. Similarly, rhBMP‐7 did not ameliorate the significant increase in urinary concentrations of transforming growth factor‐β. In summary, our in vitro data demonstrated a poor beneficial effect of rhBMP‐7 to reverse cell toxicity while, in vivo, there was no beneficial effect of rhBMP‐7. Therefore, further investigations are needed to confirm the exact role of BMP‐7 in progressive chronic kidney disease. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-07T01:38:35.977963-05:
      DOI: 10.1002/jat.3116
  • Bisphenol A promotes X‐linked inhibitor of apoptosis
           protein‐dependent angiogenesis via G protein‐coupled estrogen
           receptor pathway
    • Authors: Jian Liu; Xin Jin, Nana Zhao, Xiaolei Ye, Chenjiang Ying
      Pages: n/a - n/a
      Abstract: Bisphenol A (BPA), one of the high‐volume chemicals worldwide, has a core structure resembling that of natural estradiol. Recent evidence has demonstrated that exposure to BPA has a relationship with the risk of cancer. The objective of our study is to investigate the mechanisms underlying the pro‐angiogenic effects of BPA. We demonstrated that BPA markedly induces endothelial cell proliferation, migration and tube formation by activating endothelial nitric oxide synthase. BPA‐induced nitric oxide generation appeared to be associated with the X‐linked inhibitor of apoptosis protein (XIAP), which competes with endothelial nitric oxide synthase for caveolin‐1. BPA was shown to exert its pro‐angiogenic effect by upregulating XIAP expression via G protein‐coupled estrogen receptor (ER) activation but not via ERα or ERβ. Our data suggest that 100 nM BPA promote angiogenesis in a G protein‐coupled ER‐dependent genomic pathway, and provide a novel insight into the potential role of XIAP in mediating the pro‐angiogenic effects of BPA in endothelial cells. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-07T01:22:35.541079-05:
      DOI: 10.1002/jat.3112
  • Safety of Chinese herbal medicines during pregnancy
    • Authors: Bo Liang; Lu Li, Ling Yin Tang, Qi Wu, Xiao Ke Wu, Chi Chiu Wang
      Pages: n/a - n/a
      Abstract: Miscarriage and infertility have long been public concerns due to the mental and physical suffering they bring to potential parents. There is a strong need for effective and affordable treatments. Chinese herbal medicines (CHMs) have been shown to be effective for preventing miscarriage and treating infertility; however, due to the limited knowledge of their pharmacological mechanisms and unknown potential toxicity, their use has been restricted. This paper reviews 24 clinical trials of CHMs to prevent miscarriage and treat infertility. Most of these studies did not meet the requirements of randomized controlled trials. Even when using quality assessments based on the Newcastle–Ottawa Scale to assess the quality of non‐randomized studies, most studies did not meet the requirements. The reviewed papers were evaluated for maternal and embryonic adverse effects, including those in animal experiments. Slight maternal effects were noted, with some reports of severe toxic effects of CHMs for preventing miscarriage and severe adverse maternal effects of CHMs used for infertility. Owing to the poor quality of the randomized controlled clinical trials and the limited number of studies, it is not possible to draw a conclusion. From animal studies, for all three gestational periods, growth delay and congenital anomalies were the most commonly recorded adverse effects. However, baseline toxicological data and detailed mechanisms are still lacking. To gain a better understanding of the potential toxic effects of CHMs, additional high‐quality randomized controlled trials should be conducted, and high‐throughput in vitro screening method for baseline data should be considered. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-07T01:01:15.29916-05:0
      DOI: 10.1002/jat.3108
  • Safety assessment of aditoprim acute, subchronic toxicity and mutagenicity
    • Authors: Xu Wang; Ziqiang Tan, Yuanhu Pan, Awais Ihsan, Qianying Liu, Lingli Huang, Guyue Cheng, Dongmei Chen, Yanfei Tao, Zhenli Liu, Zonghui Yuan
      Pages: n/a - n/a
      Abstract: Aditoprim (ADP), a new developed dihydrofolate reductase (DHFR) inhibitor, has great potential in clinical veterinary medicine because of its greater pharmacokinetic properties than structural analogs. Preclinical toxicology studies were performed to assess the safety of ADP including an acute oral toxicity test, a subchronic toxicity test and five mutagenicity tests. In the acute oral toxicity test, ADP was administered singly by oral gavage to Wistar rats and Kunming mice. The LD50 calculated was 1400 mg kg–1 body weight (BW) day–1 in rats and 1130 mg kg–1 BW day–1 in mice. In a subchronic study, Wistar rats were administered ADP at dose levels of 0, 20, 100 and 1000 mg kg–1 diet for 90 days. Significant decreases were observed on body weight and food efficiency in the high‐dose group. Treatment‐related changes in clinical serum biochemistry were found in the medium‐ and high‐dose groups. Significant increases in the relative weights of livers and kidneys in females and testis in males in the 1000 mg kg–1 diet, and significant decrease in relative weights of livers in males in the 100 mg kg–1 diet were noted. Histopathological observations revealed that the 1000 mg kg–1 ADP diet could induce lymphocytic infiltration and hepatocytic necrosis near the hepatic portal area. The genotoxicity of ADP was negative in tests, such as the bacterial reverse mutation assay, mice bone marrow erythrocyte micronucleus assay, in vitro chromosomal aberration test, in vitro cho/hgprt mammalian cell mutagenesis assay and mice testicle cells chromosome aberration. Based on the subchronic study, the no‐observed‐adverse‐effect level for ADP was a 20 mg kg–1 diet, which is about 1.44‐1.53 mg kg–1 BW day–1 in rats. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-07T00:21:09.972171-05:
      DOI: 10.1002/jat.3107
  • Comparative cytotoxicity of dolomite nanoparticles in human larynx HEp2
           and liver HepG2 cells
    • Authors: Maqusood Ahamed; Hisham A. Alhadlaq, Javed Ahmad, Maqsood A. Siddiqui, Shams T. Khan, Javed Musarrat, Abdulaziz A. Al‐Khedhairy
      Pages: n/a - n/a
      Abstract: Dolomite is a natural mineral of great industrial and commercial importance. With the advent of nanotechnology, natural minerals including dolomite in the form of nanoparticles (NPs) are being utilized in various applications to improve the quality of products. However, safety or toxicity information of dolomite NPs is largely lacking. This study evaluated the cytotoxicity of dolomite NPs in two widely used in vitro cell culture models: human airway epithelial (HEp2) and human liver (HepG2) cells. Concentration‐dependent decreased cell viability and damaged cell membrane integrity revealed the cytotoxicity of dolomite NPs. We further observed that dolomite NPs induce oxidative stress in a concentration‐dependent manner, as indicated by depletion of glutathione and induction of reactive oxygen species (ROS) and lipid peroxidation. Quantitative real‐time PCR data demonstrated that the mRNA level of tumor suppressor gene p53 and apoptotic genes (bax, CASP3 and CASP9) were up‐regulated whereas the anti‐apoptotic gene bcl‐2 was down‐regulated in HEp2 and HepG2 cells exposed to dolomite NPs. Moreover, the activity of apoptotic enzymes (caspase‐3 and caspase‐9) was also higher in both kinds of cells treated with dolomite NPs. It is also worth mentioning that HEp2 cells seem to be marginally more susceptible to dolomite NPs exposure than HepG2 cells. Cytotoxicity induced by dolomite NPs was efficiently prevented by N‐acetyl cysteine treatment, which suggests that oxidative stress is primarily responsible for the cytotoxicity of dolomite NPs in both HEp2 and HepG2 cells. Toxicity mechanisms of dolomite NPs warrant further investigations at the in vivo level. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-06T23:47:14.312666-05:
      DOI: 10.1002/jat.3097
  • Are zebrafish larvae suitable for assessing the hepatotoxicity potential
           of drug candidates?
    • Authors: Natalie Mesens; Alexander D. Crawford, Aswin Menke, Pham Duc Hung, Freddy Van Goethem, Rik Nuyts, Erik Hansen, Andre Wolterbeek, Jacky Van Gompel, Peter De Witte, Camila V. Esguerra
      Pages: n/a - n/a
      Abstract: Drug‐induced liver injury (DILI) is poorly predicted by single‐cell‐based assays, probably because of the lack of physiological interactions with other cells within the liver. An intact whole liver system such as one present in zebrafish larvae could provide added value in a screening strategy for DILI; however, the possible occurrence of other organ toxicities and the immature larval stage of the zebrafish might complicate accurate and fast analysis. We investigated whether expression analysis of liver‐specific fatty acid binding protein 10a (lfabp10a) was an appropriate endpoint for assessing hepatotoxic effects in zebrafish larvae. It was found that expression analysis of lfabp10a was a valid marker, as after treatment with hepatotoxicants, dose–response curves could be obtained and statistically significant abnormal lfabp10 expression levels correlated with hepatocellular histopathological changes in the liver. However, toxicity in other vital organs such as the heart could impact liver outgrowth and thus had to be assessed concurrently. Whether zebrafish larvae were suitable for assessing human relevant drug‐induced hepatotoxicity was assessed with hepatotoxicants and non‐hepatotoxicants that have been marketed for human use and classified according to their mechanism of toxicity. The zebrafish larva showed promising predictivity towards a number of mechanisms and was capable of distinguishing between hepatotoxic and non‐hepatotoxic chemical analogues, thus implying its applicability as a potential screening model for DILI. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-06T22:52:02.514727-05:
      DOI: 10.1002/jat.3091
  • Bupropion treatment increases epididymal contractility and impairs sperm
    • Authors: Marilia Martins Cavariani; Luiz Ricardo Almeida Kiguti, Josiane Lima Rosa, Gabriel Adan Araújo Leite, Patrícia Villela e Silva, André Sampaio Pupo, Wilma De Grava Kempinas
      Abstract: Bupropion is a dopamine (DA) and norepinephrine (NE) reuptake inhibitor used as smoking cessation and antidepressant drug with a lower incidence of male sexual dysfunction. We showed previously that sibutramine, a norepinephrine/serotonine reuptake inhibitor, reduced male rat fertility. As there are no studies evaluating the impact of bupropion treatment on spermatic parameters and male fertility, we evaluated the effects of bupropion treatment (15 and 30 mg kg−1, 30 days) on sexual behavior, spermatic parameters and fertility of male Wistar rats and on the epididymal duct in vitro contractility. Bupropion 15 mg kg−1 increased the serum luteinizing hormone level and the epididymal duct contractility, but the sperm quality was not affected. At 30 mg kg−1 bupropion impaired sperm quality increasing the incidence of non‐progressive sperm. The male sexual behavior and fertility were not modified at both bupropion doses. These results, in rats, suggest the importance of studies evaluating the effects of bupropion on the human male sperm quality. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-02T20:20:56.934592-05:
      DOI: 10.1002/jat.3089
  • Molecular mechanisms of human thyrocyte dysfunction induced by low
           concentrations of polychlorinated biphenyl 118 through the Akt/FoxO3a/NIS
    • Authors: Hongwei Guo; Hui Yang, Huanhuan Chen, Wen Li, Jinmei Tang, Pei Cheng, Yuchun Xie, Yun Liu, Guoxian Ding, Dai Cui, Xuqin Zheng, Yu Duan
      Abstract: Polychlorinated biphenyls (PCBs) are typical persistent organic pollutants that can interfere with multiple organ systems of humans. Previously, we concluded that persistent exposure to low doses of PCB118 could severely damage the thyroidal structure, dramatically decrease the concentration of serum thyroid hormones and inhibit the pivotal gene expressions such as sodium/iodide symporter (NIS) and thyroglobulin (Tg). To explore the molecular mechanisms of thyrocyte dysfunction induced by 2,3′,4,4′,5‐pentachlorobiphenyl (PCB118), monolayer cultured human thyroid epithelial cells (HTECs) were treated with PCB118 or dimethyl sulfoxide (DMSO) as a control. Our results indicated that relatively higher concentrations of PCB118 could induce a loss in the viability of HTEC. In cultures with concentrations of PCB118 from 0.025 to 25 nM, which did not affect cell viability or apoptosis, concentrations of Tg and thyroxine (T4) were significantly decreased compared with those in the controls. In addition, mRNA and protein levels of Akt were increased significantly in the PCB118‐treated groups, whereas FoxO3a expression did not show particular variation. Furthermore, exposure to PCB118 was associated with a significant increase of the protein levels of p‐Akt and p‐FoxO3a, and these effects were blocked by LY294002. In contrast, mRNA and protein expression levels of NIS were decreased significantly, and this effect was blocked by LY294002. Unlike control cells, a cytoplasmic shift of FoxO3a was observed in the PCB118‐treated group. Our research suggests that PCB118 may induce thyrocyte dysfunction through the Akt/FoxO3a/NIS signalling pathway, which provides potential new insights for finding interventions to counteract the damage to the human body caused by PCBs. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-02T19:57:38.986114-05:
      DOI: 10.1002/jat.3032
  • Different cytotoxicity responses to antimicrobial nanosilver coatings when
           comparing extract‐based and direct‐contact assays
    • Authors: Eric M. Sussman; Brendan J. Casey, Debargh Dutta, Benita J. Dair
      Abstract: This study was performed to understand how the choice of cytotoxicity assay format affects the observed biocompatibility of nanosilver (nAg). nAg coatings are physical coatings containing silver (Ag) that have feature sizes of 100 nm or less, often in the form of nanoparticles or grains. They are used on medical devices to prevent infection, but in spite of this intended benefit, observations of potential cytotoxicity from nAg have been reported in numerous published studies. For medical device regulation, cytotoxicity testing is part of a biocompatibility evaluation, in which specific test methods are chosen based on the technological characteristics and intended use of a device. For this study, nAg‐coated tissue culture polystyrene surfaces were prepared using magnetron sputter coating, resulting in nAg films of 0.2 to 311 µg cm−2 Ag. These coatings exhibited nanometer‐scale morphologies and demonstrated a > 4log10 reduction in Escherichia coli viability. It was observed that extracts of nAg caused no cytotoxicity to L929 mouse fibroblasts, but cells cultured directly on nAg coatings (direct‐contact assay format) showed a dose‐dependent reduction in viability by up to 100% (P 
      PubDate: 2015-02-02T18:51:12.25067-05:0
      DOI: 10.1002/jat.3104
  • Effects of cylindrospermopsin on the phagocytic cells of the common carp
           (Cyprinus carpio L.)
    • Authors: Anna Sieroslawska; Anna Rymuszka, Łukasz Adaszek
      Abstract: Cylindrospermopsin is a cyanotoxin with cytotoxic activity. It is released into water during and after cyanobacterial water blooms and thus poses a threat to the health of fish. There is very little information available concerning the effects of the toxin on fish immune cells. In this study, we assessed the potential impact of cylindrospermopsin on the basic functions of phagocytic cells from common carp (Cyprinus carpio L.), including phagocytosis, reactive oxygen and nitrogen species production, and the structure of microfilaments and selected cytokine expression. Phagocytic cells, isolated from fish head kidneys, were exposed to the toxin at concentrations of 0.05, 0.1, 0.5 or 1 µg ml−1, for up to 24 h. Cytotoxicity, detected by lactate dehydrogenase release, was observed at the highest studied concentration. A decrease in phagocytic activity and changes in actin cytoskeletal structures were observed after the cell exposure to the toxin at 0.5 and 1 µg ml−1. Moreover, at all tested concentrations, cylindrospermopsin increased the production of reactive oxygen and nitrogen species. It also evidently influenced the expression of genes of proinflammatory cytokines interleukin‐1β and tumour necrosis factor‐α and, to a minor extent, anti‐inflammatory transforming growth factor‐β, but had no effects on interleukin‐10. The results indicated that the cyanotoxin cylindrospermopsin is able to modify basic features of carp phagocytic cells, which might result in adverse consequences for fish health. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-29T08:58:37.744329-05:
      DOI: 10.1002/jat.3118
  • The relationship between Cd‐induced autophagy and lysosomal
           activation in WRL‐68 cells
    • Authors: Su‐Fang Meng; Wei‐Ping Mao, Fang Wang, Xiao‐Qian Liu, Luan‐Luan Shao
      Abstract: This study shows that Cd induces autophagy in the human's embryonic normal liver cell line (WRL‐68). The expression of LC3B‐II and the mature cathepsin L were analyzed by Western blotting. The autophagosomes and lysosomes were directly visualized by electron microscopy and confocal microscopy analysis in Cd‐exposed WRL‐68 cells. In this study, we first found that autophagy induced the activation of lysosomal function in WRL‐68 cells. The lysosomal activation was markedly decreased when the cells were co‐treated with 3‐MA (an inhibitor of autophagy). Secondly, we provided the evidence that the activation of lysosomal function depended on autophagosome–lysosome fusion. The colocalization of lysosome‐associated membrane protein‐2 (LAMP2) and GFP‐LC3 was significantly reduced, when they were treated with thapsigargin (an inhibitor of autophagosome–lysosome fusion). We demonstrated that deletion or blockage of the autophagosome–lysosome fusion process effectively diminished lysosomal activation, which suggests that lysosomal activation occurring in the course of autophagy is dependent on autophagosome–lysosome fusion. Thirdly, we provided evidence that the activation of lysosomal function was associated with lysosomal acid. We investigated the relationship between autophagosome–lysosome fusion and pH in acidic compartments by visualizing fusion process in WRL‐68 cells. This suggests that increasing pH in acidic compartments in WRL‐68 cells inhibits the autophagosome–lysosome fusion. Finally, we found that the activation of lysosomal function was associated with Ca2+ stores and the intracellular Ca2+ channels or pumps were possibly pH‐dependent. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-29T08:46:15.013806-05:
      DOI: 10.1002/jat.3114
  • Bisphenol A exposure induces metabolic disorders and enhances
           atherosclerosis in hyperlipidemic rabbits
    • Authors: Chao Fang; Bo Ning, Ahmed Bilal Waqar, Manabu Niimi, Shen Li, Kaneo Satoh, Masashi Shiomi, Ting Ye, Sijun Dong, Jianglin Fan
      Abstract: Bisphenol A (BPA) is an artificial environmental endocrine disrupter. Excess exposure to BPA may induce many disorders in the metabolism and cardiovascular system. However, the underlying toxicological mechanisms remain largely unknown. In this study, we administered genetically hyperlipidemic Watanabe heritable hyperlipidemic (WHHL‐MI) rabbits (male, 14 week old), which have more common features with humans than the mouse and rat especially in the metabolism and cardiovascular system, with BPA at 40 mg kg–1 day–1 for 8 weeks by gavage and compared their plasma lipids, glucose and insulin response with those of the vehicle group. All of the rabbits were sacrificed, and their pancreas, liver, adipose tissue, heart and aorta were analyzed using histological and morphometric methods. Furthermore, we treated human hepatoma HepG2 cells and human umbilical cord vein endothelial cells (HUVECs), with different doses of BPA based on the serum BPA levels in the WHHL rabbits for 6 h to investigate the possible molecular mechanisms. Our results showed that BPA‐treated rabbits showed insulin resistance, prominent adipose accumulation and hepatic steatosis. Additionally, BPA exposure also caused myocardial injury and enhanced the development of atherosclerosis in the aortic arch with increased macrophage number (86%) and advanced lesion areas (69%). Increased expression of inflammatory genes found in the liver of BPA‐treated rabbits along with the up‐regulation of ER stress, lipid and glucose homeostasis and inflammatory genes in the cultured HepG2 cells and HUVECs suggest that BPA may induce metabolic disorders and enhance atherosclerosis through regulating above molecular pathways in the liver and endothelium. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-23T13:24:22.578133-05:
      DOI: 10.1002/jat.3103
  • Effects of homocysteine on mesenchymal cell proliferation and
           differentiation during chondrogenesis on limb development
    • Authors: Gilian Fernando Bourckhardt; Manuela Sozo Cecchini, Dib Ammar, Karoline Kobus‐Bianchini, Yara Maria Rauh Müller, Evelise Maria Nazari
      Abstract: High levels of homocysteine (Hcy) are related to an increased risk of the occurrence of congenital anomalies, including limb defects. However, few evaluations about how toxic levels of Hcy affect limb development have been reported. We investigated whether Hcy can affect the cell cycle proteins and proteins involved in mesenchymal cell differentiation during limb development, in a chicken embryo model. Embryos were treated with 20 µmol d‐l Hcy/50 µl saline at embryonic day 2 and analyzed at embryonic day 6. Untreated control embryos received exclusively 50 µl saline solution. To identify cells in proliferation and cell cycle proteins, as well as Pax1/9 and Sox9 proteins, we performed immunolocalization and flow cytometry analyses using the antibodies anti‐phosphohistone H3, anti‐p53, anti‐p21, anti‐proliferating cell nuclear antigen, anti‐Pax1, anti‐Pax9 and anti‐Sox9. No significant differences in cell proliferation were observed between Hcy‐treated and untreated embryos. We observed a decrease of the proliferating cell nuclear antigen and p21 proteins, both involved in the G1 phase of cell cycle progression. On the other hand, in mesenchymal cells of the limbs, Hcy induces an increase of p53 protein, which can be activated by DNA damage. In cell differentiation, Hcy induced an increase mainly of Pax9 and Sox9 proteins. Our data indicate that the treatment with Hcy changes the mesenchymal cell dynamics during limb development, but does not change the morphology of the cartilage molds. These findings provide information to understand better the cellular basis of the toxicity of Hcy on chondrogenesis during limb development. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-23T12:15:56.210643-05:
      DOI: 10.1002/jat.3111
  • Quantitative evaluation of the pulmonary microdistribution of TiO2
           nanoparticles using X‐ray fluorescence microscopy after
           intratracheal administration with a microsprayer in rats
    • Authors: Guihua Zhang; Naohide Shinohara, Hirokazu Kano, Hideki Senoh, Masaaki Suzuki, Takeshi Sasaki, Shoji Fukushima, Masashi Gamo
      Abstract: The unevenness of pulmonary nanoparticle (NP) distribution, which hinders the establishment of an absolute dose–response relationship, has been described as one of the limitations of intratracheal administration techniques for toxicological assessment of inhaled NPs. Quantification of the NP microdistribution would facilitate the establishment of a concentration–response relationship in localized regions of the lung; however, such quantitative methods have not been reported. Here, we established a quantitative method for evaluating pulmonary TiO2 NP microdistribution in rats using X‐ray fluorescence microscopy. Ti intensity in lung sections from rats intratracheally administered 10 mg kg–1 TiO2 NPs with a microsprayer was measured using X‐ray fluorescence with a 100 µm beam size. Ti reference samples were prepared by dropping different concentrations of Ti solutions on glass slide or lung sections of untreated rat. Ti intensity increased linearly with Ti content in the reference samples on both substrates. The detection limit of TiO2 was estimated to be 6.3 ng mm–2. The reproducibility was confirmed for measurements done in the short‐ (2 weeks) and long‐term (6 months). The quantitative results of TiO2 NP microdistribution suggested that more TiO2 NPs were distributed in the right caudal and accessory lobes, which are located downstream of the administration direction of the NP suspension, and the lower portion of each lobe. The detection rates of TiO2 NPs were 16.6–25.0%, 5.19–15.6%, 28.6–39.2%, 21.4–38.7% and 10.6–23.2% for lung sections from the right cranial, middle, caudal, accessory and left lobes, respectively. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-23T11:49:20.327102-05:
      DOI: 10.1002/jat.3109
  • Endocrine‐disrupting potentials of equine estrogens equilin,
           equilenin, and their metabolites, in the medaka Oryzias latipes: in silico
           and DNA microarray studies
    • Authors: Masaya Uchida; Hiroshi Ishibashi, Ryoko Yamamoto, Akiko Koyanagi, Teruhiko Kusano, Nobuaki Tominaga, Yasuhiro Ishibashi, Koji Arizono
      Abstract: Although several previous studies have demonstrated the presence of equine estrogens in the aquatic environment, limited data are currently available on the endocrine‐disrupting potentials in fish and the risks they pose to aquatic organisms. To investigate the interactions of major equine estrogens equilin (Eq) and equilenin (Eqn), as well as their metabolites 17α‐dihydroequilin, 17β‐dihydroequilin, 17α‐dihydroequilenin and 17β‐dihydroequilenin, with the estrogen receptor α (ERα) of medaka (Oryzias latipes), a three‐dimensional model of the ligand‐binding domain (LBD) of ERα was built in silico, and docking simulations were performed. The docking simulation analysis indicated that the interaction of 17β‐dihydroequilenin with the ERα LBD is the most potent, followed by those of 17α‐dihydroequilin and 17β‐dihydroequilin, whereas those of Eq and Eqn were least potent. We further analyzed gene expression profiles in the livers of male medaka exposed to Eq and Eqn. A DNA microarray representing 6000 genes revealed that 24‐h exposure to Eq and Eqn (100 ng/L) upregulated the expression of 6 and 34 genes in the livers of males, respectively. Genes upregulated by Eq included the estrogenic biomarker genes vitellogenins and choriogenins, suggesting the estrogenic potential of Eq. In contrast, Eqn exposure upregulated several cancer‐related genes, such as mediator complex subunit 16 and RAS oncogene family members, suggesting a carcinogenic potential for Eqn. These results suggest that equine estrogens may have not only endocrine‐disrupting potentials via the ERα signaling pathway but also carcinogenic potency in male medaka. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-22T00:34:36.505314-05:
      DOI: 10.1002/jat.3098
  • Effects of lithium on growth, maturation, reproduction and gene expression
           in the nematode Caenorhabditis elegans
    • Authors: Ayako Inokuchi; Ryoko Yamamoto, Fumiyo Morita, Shota Takumi, Hiromi Matsusaki, Hiroshi Ishibashi, Nobuaki Tominaga, Koji Arizono
      Abstract: Lithium (Li) has been widely used to treat bipolar disorder, and industrial use of Li has been increasing; thus, environmental pollution and ecological impacts of Li have become a concern. This study was conducted to clarify the potential biological effects of LiCl and Li2CO3 on a nematode, Caenorhabditis elegans as a model system for evaluating soil contaminated with Li. Exposure of C. elegans to LiCl and Li2CO3 decreased growth/maturation and reproduction. The lowest observed effect concentrations for growth, maturation and reproduction were 1250, 313 and 10 000 µm, respectively, for LiCl and 750, 750 and 3000 µm, respectively, for Li2CO3. We also investigated the physiological function of LiCl and LiCO3 in C. elegans using DNA microarray analysis as an eco‐toxicogenomic approach. Among approximately 300 unique genes, including metabolic genes, the exposure to 78 µm LiCl downregulated the expression of 36 cytochrome P450, 16 ABC transporter, 10 glutathione S‐transferase, 16 lipid metabolism and two vitellogenin genes. On the other hand, exposure to 375 µm Li2CO3 downregulated the expression of 11 cytochrome P450, 13 ABC transporter, 13 lipid metabolism and one vitellogenin genes. No gene was upregulated by LiCl or Li2CO3. These results suggest that LiCl and Li2CO3 potentially affect the biological and physiological function in C. elegans associated with alteration of the gene expression such as metabolic genes. Our data also provide experimental support for the utility of toxicogenomics by integrating gene expression profiling into a toxicological study of an environmentally important organism such as C. elegans. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-21T22:52:27.326537-05:
      DOI: 10.1002/jat.3058
  • Cytotoxicity of luteolin in primary rat hepatocytes: the role of
           CYP3A‐mediated ortho‐benzoquinone metabolite formation and
           glutathione depletion
    • Authors: Fuguo Shi; Peng Zhao, Xiaobing Li, Hong Pan, Shiping Ma, Li Ding
      Abstract: Luteolin (LUT), an active ingredient in traditional Chinese medicines and an integral part of the human diet, has shown promising pharmacological activities with a great potential for clinical use. The purpose of this study was to evaluate the role of cytochrome P450 (CYP450)‐mediated reactive ortho‐benzoquinone metabolites formation and glutathione (GSH) depletion in LUT‐induced cytotoxicity in primary rat hepatocytes. A reactive ortho‐benzoquinone metabolite was identified by liquid chromatography coupled with tandem mass spectrometry (LC‐MS/MS) in rat liver microsomes (RLMs) and rat hepatocytes. Using a specific chemical inhibitor method, the CYP3A subfamily was found to be responsible for the reactive metabolite formation in RLMs. Induction of CYP3A by dexamethasone enhanced LUT‐induced cytotoxicity, whereas inhibition of CYP3A by ketoconazole (Keto) decreased the cytotoxicity. The cytotoxicity and cell apoptosis induced by LUT were related to the amount of reactive metabolite formation. Furthermore, Keto inhibited the LUT‐induced GSH exhaustion. The cytotoxicity was significantly enhanced by pretreatment with L‐buthionine sulfoximine to deplete the intracellular GSH. A time course experiment showed that GSH depletion by LUT was not via oxidation of GSH and occurred prior to the increase in 2', 7'‐dichlorofluorescein in hepatocytes. Collectively, these data suggest that CYP3A‐mediated reactive metabolite formation plays a critical role in LUT‐induced hepatotoxicity, and the direct GSH depletion is an initiating event in LUT‐mediated cytotoxicity in primary rat hepatocytes. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-21T22:30:19.0298-05:00
      DOI: 10.1002/jat.3106
  • Genomic and gene expression responses to genotoxic stress in PAC2
           zebrafish embryonic cell line
    • Authors: Maja Šrut; Jean‐Paul Bourdineaud, Anamaria Štambuk, Göran I. V. Klobučar
      Abstract: PAC2 cell line is, along most of the developed zebrafish cell lines, poorly characterized concerning its response to genotoxicants. To define the PAC2 cell line response to different forms of genotoxic stress, we exposed the cells to model genotoxic agents (benzo[a]pyrene, B[a]P, and ethyl methanesulfonate) and subsequently monitored DNA damage and alterations by using the battery of tests, including the Comet assay, quantitative random‐amplified polymorphic DNA and amplified fragment length polymorphism. The expression of several DNA repair (xpc, xpd, hr23b, rad51, msh2) and oxidative stress response (sod (Cu/Zn)) genes was monitored as well. To obtain an indication of the PAC2 cell line metabolizing capacity, the expression of genes belonging to cyp1, cyp2 and cyp3 families was assessed upon exposure to B[a]P. Genotoxic responses were observed in all the used methods, and quantitative random‐amplified polymorphic DNA and amplified fragment length polymorphism proved to be more sensitive by revealing DNA alterations even when the Comet assay indicated lack of significant damage. The PAC2 cell line demonstrated basal and B[a]P‐induced expression of several cyp genes, suggesting its ability to metabolize indirect acting xenobiotics to a certain point. Based on these results, PAC2 cells seem to be sensitive zebrafish in vitro model in the genotoxicity assessment of the direct acting genotoxicant; however, they are less sensitive toward the indirect acting genotoxicant due to their limited metabolizing properties. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-21T22:16:15.203856-05:
      DOI: 10.1002/jat.3113
  • Toxicity of cobalt–chromium nanoparticles released from a
           resurfacing hip implant and cobalt ions on primary human lymphocytes in
    • Authors: Olga M. Posada; R. J. Tate, M. H. Grant
      Abstract: Adverse tissue responses to prostheses wear particles and released ions are important contributors to hip implant failure. In implant‐related adverse reactions T‐lymphocytes play a prominent role in sustaining the chronic inflammatory response. To further understand the involvement of lymphocytes in metal‐on‐metal (MoM) implant failure, primary human lymphocytes were isolated and treated with cobalt–chromium (Co‐Cr) wear debris and Co ions, individually, and in combination, for 24, 48 and 120 h. There was a significant increase in cell number where debris was present, as measured by the Neutral Red assay. Interleukin‐6 (IL‐6), interferon‐γ (IFN‐γ) and tumour necrosis factor‐α (TNF‐α) secretion levels significantly decreased in the presence of metal particles, as measured by ELISA. Interleukin‐2 (IL‐2) secretion levels were significantly decreased by both debris and Co ions. Flow cytometry analysis showed that the metal nanoparticles induced a significant increase in apoptosis after 48‐h exposure. This investigation showed that prolonged exposure (120 h) to metal debris induces lymphocyte proliferation, suggesting that activation of resting lymphocytes may have occurred. Although cytokine production was affected mainly by metal debris, cobalt toxicity may also modulate IL‐2 secretion, and even Co ion concentrations below the MHRA guideline levels (7 ppb) may contribute to the impairment of immune regulation in vivo in patients with MoM implants. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-21T21:31:14.028782-05:
      DOI: 10.1002/jat.3100
  • Cobalt oxide nanoparticles induced oxidative stress linked to activation
           of TNF‐α/caspase‐8/p38‐MAPK signaling in human
           leukemia cells
    • Authors: Sourav Chattopadhyay; Sandeep Kumar Dash, Satyajit Tripathy, Balaram Das, Santanu Kar Mahapatra, Panchanan Pramanik, Somenath Roy
      Abstract: The purpose of this study was to determine the intracellular signaling transduction pathways involved in oxidative stress induced by nanoparticles in cancer cells. Activation of reactive oxygen species (ROS) has some therapeutic benefits in arresting the growth of cancer cells. Cobalt oxide nanoparticles (CoO NPs) are an interesting compound for oxidative cancer therapy. Our results showed that CoO NPs elicited a significant (P
      PubDate: 2015-01-11T19:52:09.170136-05:
      DOI: 10.1002/jat.3080
  • Quantitative toxicoproteomic analysis of zebrafish embryos exposed to a
           retinoid X receptor antagonist UVI3003
    • Authors: Liang Zheng; Jianlan Yu, Huahong Shi, Liang Xia, Qi Xin, Qiang Zhang, Heng Zhao, Ji Luo, Wenhai Jin, Daoji Li, Junliang Zhou
      Abstract: Retinoid X receptor (RXR) antagonists, including some environmental endocrine disruptors, have a teratogenic effect on vertebrate embryos. To investigate the toxicological mechanism on the protein expression level, a quantitative proteomic study was conducted to analyze the proteome alterations of zebrafish (Danio rerio) embryos exposed to gradient concentrations of a representative RXR antagonist UVI3003. Using isobaric Tags for Relative and Absolute Quantitation (iTRAQ) labeling coupled nano high‐performance liquid chromatography‐tandem mass spectrometry (nano HPLC‐MS/MS), in total 6592 proteins were identified, among which 195 proteins were found to be differentially expressed by more than a two‐fold change in exposed groups compared with the control. Gene ontology analysis showed that these differential proteins were mostly involved in anatomical structure development, biosynthetic process, ion binding and oxidoreductase activity. Moreover, the biological pathways of translation, lipoprotein metabolism, cell survival and gluconeogenesis were intensively inhibited after exposure. Some significantly downregulated proteins such as apolipoprotein A‐I and vitellogenin and upregulated proteins such as calcium activated nucleotidase 1b, glutathione S‐transferase and glucose 6‐dehydrogenases showed a strong dose‐dependent response. The results provided new insight into the molecular details of RXR antagonist‐induced teratogenicity and added novel information of pathways and potential biomarkers for evaluation of RXR interfering activity. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-11T19:41:31.661922-05:
      DOI: 10.1002/jat.3099
  • The toxicity and distribution of iron oxide–zinc oxide
           core‐shell nanoparticles in C57BL/6 mice after repeated subcutaneous
    • Authors: Jun‐Won Yun; Jung‐Hee Yoon, Byeong‐Cheol Kang, Nam‐Hyuk Cho, Seung Hyeok Seok, Seung‐Kee Min, Ji Hyun Min, Jeong‐Hwan Che, Young Keun Kim
      Abstract: Therapeutic cancer vaccines promote immune responses by delivering tumour‐specific antigens. Recently, we developed iron oxide (Fe3O4)–zinc oxide (ZnO) core‐shell nanoparticles (CSNPs) as carriers for antigen delivery into dendritic cells (DCs), and the CSNPs were injected subcutaneously into C57BL/6 mice to examine the systemic toxicity, tissue distribution and excretion of the CSNPs. The doses injected were 0, 4, 20 and 200 mg kg–1 weekly for 4 weeks. No significant changes were observed after the CSNPs administration with respect to mortality, clinical observations, body weight, food intake, water consumption, urinalysis, haematology, serum biochemistry,and organ weights. A dose‐dependent increase in granulomatous inflammation was observed at the injection site of the CSNP‐treated animals, but no other histopathological lesions in other organs could be attributed to the CSNPs. The Zn concentration, which is an indicator for CSNPs, was not significantly higher in the sampled tissues, urine, or faeces after the CSNP injection. In contrast, the Zn concentration at the subcutaneous skin of the site injected with the CSNPs increased in a dose‐dependent manner, along with a macroscopic deposition of the CSNPs. The CSNP residue at the injection site resulted in a foreign body response with the appearance of macrophage infiltration, but otherwise did not show any systemic distribution or toxicity at up to 200 mg kg–1 during this study. In conclusion, CSNPs could be used as good antigen carriers for DC‐based immunotherapy, although further study is needed to completely clear the residue of the CSNPs at the injection site. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-08T20:58:03.730817-05:
      DOI: 10.1002/jat.3102
  • Enhanced QSAR models for drug‐triggered inhibition of the main
           cardiac ion currents
    • Authors: Barbara Wiśniowska; Aleksander Mendyk, Jakub Szlęk, Michał Kołaczkowski, Sebastian Polak
      Abstract: The currently changing cardiac safety testing paradigm suggests, among other things, a shift towards using in silico models of cellular electrophysiology and assessment of a concomitant block of multiple ion channels. In this study, a set of four enhanced QSAR models have been developed: for the rapid delayed rectifying potassium current (IKr), slow delayed rectifying potassium current (IKs), peak sodium current (INa) and late calcium current (ICaL), predicting ion currents changes for the specific in vitro experiment from the 2D structure of the compounds. The models are a combination of both in vitro study parameters and physico‐chemical descriptors, which is a novel approach in drug–ion channels interactions modeling. Their predictive power assessed in the enhanced, more demanding than standard procedure, 10‐fold cross validation was reasonably high. Rough comparison with published pure in silico hERG interaction models shows that the quality of the model predictions does not differ from other models available in the public domain, however, it takes its advantage in accounting for inter‐experimental settings variability. Developed models are implemented in the Cardiac Safety Simulator, a commercially available platform enabling the in vitro–in vivo extrapolation of the drugs proarrhythmic effect and ECG simulation. A more comprehensive assessment of the effects of the compounds on ion channels allows for making more informed decisions regarding the risk – and thus avoidance – of exclusion of potentially safe and effective drugs. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-01-05T20:26:17.842509-05:
      DOI: 10.1002/jat.3095
  • Tl(I) and Tl(III) alter the expression of EGF‐dependent signals and
           cyclins required for pheochromocytoma (PC12) cell‐cycle resumption
           and progression
    • Authors: María T. L. Pino; Sandra V. Verstraeten
      Abstract: The effects of thallium [Tl(I) and Tl(III)] on the PC12 cell cycle were evaluated without (EGF−) or with (EGF+) media supplementation with epidermal growth factor (EGF). The following markers of cell‐cycle phases were analyzed: cyclin D1 (G1); E2F‐1, cyclin E and cytosolic p21 (G1→S transition); nuclear PCNA and cyclin A (S); and cyclin B1 (G2). The amount of cells in each phase and the activation of the signaling cascade triggered by EGF were also analyzed. Tl(I) and Tl(III) (5–100 μM) caused dissimilar effects on PC12 cell proliferation. In EGF− cells, Tl(I) increased the expression of G1→S transition markers and nuclear PCNA, without affecting cyclin A or cyclin B1. In addition to those, cyclin B1 was also increased in EGF+ cells. In EGF− cells, Tl(III) increased the expression of cyclin D1, all the G1→S and S phase markers and cyclin B1. In EGF+ cells, Tl(III) increased cyclin D1 expression and decreased all the markers of G1→S transition and the S phase. Even when these cations did not induce the activation of EGF receptor (EGFR) in EGF− cells, they promoted the phosphorylation of ERK1/2 and Akt. In the presence of EGF, the cations anticipated EGFR phosphorylation without affecting the kinetics of EGF‐dependent ERK1/2 and Akt phosphorylation. Altogether, results indicate that Tl(I) promoted cell proliferation in both EGF− and EGF+ cells. In contrast, Tl(III) promoted the proliferation of EGF− cells but delayed it in EGF+ cells, which may be related to the toxic effects of this cation in PC12 cells. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-12-22T10:41:36.332836-05:
      DOI: 10.1002/jat.3096
  • All‐cause mortality increased by environmental cadmium exposure in
           the Japanese general population in cadmium non‐polluted areas
    • Authors: Yasushi Suwazono; Kazuhiro Nogawa, Yuko Morikawa, Muneko Nishijo, Etsuko Kobayashi, Teruhiko Kido, Hideaki Nakagawa, Koji Nogawa
      Abstract: The aim of the present study was to evaluate the effect of environmental cadmium (Cd) exposure indicated by urinary Cd on all‐cause mortality in the Japanese general population. A 19‐year cohort study was conducted in 1067 men and 1590 women aged 50 years or older who lived in three cadmium non‐polluted areas in Japan. The subjects were divided into four quartiles based on creatinine adjusted U‐Cd (µg g−1 cre). The hazard ratio (HR) and 95% confidence interval (CI) for continuous U‐Cd or the quartiles of U‐Cd were estimated for all‐cause mortality using a proportional hazards regression.The all‐cause mortality rates per 1000 person years were 31.2 and 15.1 in men and women, respectively. Continuous U‐Cd (+1 µg g−1 cre) was significantly related to the all‐cause mortality in men (HR 1.05, 95% CI: 1.02–1.09) and women (HR 1.04, 95% CI: 1.01–1.07). Furthermore in men, the third (1.96–3.22 µg g−1 cre) and fourth quartile (≥3.23 µg g−1 cre) of U‐Cd showed a significant, positive HR (third: HR 1.35, 95% CI: 1.03–1.77, fourth: HR 1.64, 95% CI: 1.26–2.14) for all‐cause mortality compared with the first quartile (
      PubDate: 2014-12-22T09:48:50.933751-05:
      DOI: 10.1002/jat.3077
  • Culture conditions profoundly impact phenotype in BEAS‐2B, a human
           pulmonary epithelial model
    • Authors: Fei Zhao; Walter T. Klimecki
      Pages: n/a - n/a
      Abstract: BEAS‐2B, an immortalized, human lung epithelial cell line, has been used to model pulmonary epithelial function for over 30 years. The BEAS‐2B phenotype can be modulated by culture conditions that include the presence or absence of fetal bovine serum (FBS). The popularity of BEAS‐2B as a model of arsenic toxicology, and the common use of BEAS‐2B cultured both with and without FBS, led us to investigate the impact of FBS on BEAS‐2B in the context of arsenic toxicology. Comparison of genome‐wide gene expression in BEAS‐2B cultured with or without FBS revealed altered expression in several biological pathways, including those related to carcinogenesis and energy metabolism. Real‐time measurements of oxygen consumption and glycolysis in BEAS‐2B demonstrated that FBS culture conditions were associated with a 1.4‐fold increase in total glycolytic capacity, a 1.9‐fold increase in basal respiration, a 2.0‐fold increase in oxygen consumed for ATP production and a 2.8‐fold increase in maximal respiration, compared with BEAS‐2B cultured without FBS. Comparisons of the transcriptome changes in BEAS‐2B resulting from FBS exposure to the transcriptome changes resulting from exposure to 1 μM sodium arsenite revealed that mRNA levels of 43% of the arsenite‐modulated genes were also modulated by FBS. Cytotoxicity studies revealed that BEAS‐2B cells exposed to 5% FBS for 8 weeks were almost 5 times more sensitive to arsenite cytotoxicity than non‐FBS‐exposed BEAS‐2B cells. Phenotype changes induced in BEAS‐2B by FBS suggest that culture conditions should be carefully considered when using BEAS‐2B as an experimental model of arsenic toxicity. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-12-19T00:27:35.684986-05:
      DOI: 10.1002/jat.3094
  • Ginsenoside Re protects methamphetamine‐induced mitochondrial
           burdens and proapoptosis via genetic inhibition of protein kinase C δ
           in human neuroblastoma dopaminergic SH‐SY5Y cell lines
    • Authors: Yunsung Nam; Myung Bok Wie, Eun‐Joo Shin, Thuy‐Ty Lan Nguyen, Seung‐Yeol Nah, Sung Kwon Ko, Ji Hoon Jeong, Choon‐Gon Jang, Hyoung‐Chun Kim
      Pages: n/a - n/a
      Abstract: Recently, we have demonstrated that ginsenoside Re protects methamphetamine (MA)‐induced dopaminergic toxicity in mice via genetic inhibition of PKCδ and attenuation of mitochondrial stress. In addition, we have reported that induction of mitochondrial glutathione peroxidase (GPx) is also important for neuroprotection mediated by ginsenoside Re. To extend our knowledge, we examined the effects of ginsenoside Re against MA toxicity in vitro condition using SH‐SY5Y neuroblastoma cells. Treatment with ginsenoside Re resulted in significant attenuations against a decrease in the activity of GPx and an increase in the activity of superoxide dismutase (SOD) in the cytosolic and mitochondrial fraction. The changes in glutathione (GSH) paralleled those in GPx in the same experimental condition. Consistently, ginsenoside Re treatment exhibited significant protections against cytosolic and mitochondrial oxidative damage (i.e. lipid peroxidation and protein oxidation), mitochondrial translocation of PKCδ, mitochondrial dysfunction (mitochondrial transmembrane potential and intra‐mitochondrial Ca2+), apoptotic events [i.e., cytochrome c release from mitochondria, cleavage of caspase‐3 and poly(ADP‐ribose)polymerase‐1, nuclear condensation, terminal deoxynucleotidyl transferase‐mediated dUTP nick‐end labeling (TUNEL)‐positive apoptotic cells], and a reduction in the tyrosine hydroxylase (TH) expression and TH activity induced by MA in SH‐SY5Y neuroblastoma cells. These protective effects of ginsenoside Re were comparable to those of PKCδ antisense oligonucleotide (ASO). However, ginsenoside Re did not significantly provide additional protective effects mediated by genetic inhibition of PKCδ. Our results suggest that PKCδ is a specific target for ginsenoside Re‐mediated protective activity against MA toxicity in SH‐SY5Y neuroblastoma cells. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-12-18T23:49:04.365172-05:
      DOI: 10.1002/jat.3093
  • Combined exposure to lead, inorganic mercury and methylmercury shows
           deviation from additivity for cardiovascular toxicity in rats
    • Authors: Tanja M. Wildemann; Lynn P. Weber, Steven D. Siciliano
      Pages: n/a - n/a
      Abstract: Environmental exposure to metal mixtures in the human population is common. Mixture risk assessments are often challenging because of a lack of suitable data on the relevant mixture. A growing number of studies show an association between lead or mercury exposure and cardiovascular effects. We investigated the cardiovascular effects of single metal exposure or co‐exposure to methylmercury [MeHg(I)], inorganic mercury [Hg(II)] and lead [Pb(II)]. Male Wistar rats received four different metal mixtures for 28 days through the drinking water. The ratios of the metals were based on reference and environmental exposure values. Blood and pulse pressure, cardiac output and electrical activity of the heart were selected as end‐points. While exposure to only MeHg(I) increased the systolic blood pressure and decreased cardiac output, the effects were reversed with combined exposures (antagonism). In contrast to these effects, combined exposures negatively affected the electrical activity of the heart (synergism). Thus, it appears that estimates of blood total Hg levels need to be paired with estimates of what species of mercury dominate exposure as well as whether lead co‐exposure is present to link total blood Hg levels to cardiovascular effects. Based on current human exposure data and our results, there may be an increased risk of cardiac events as a result of combined exposures to Hg(II), MeHg(I) and Pb(II). This increased risk needs to be clarified by analyzing lead and Hg exposure data in relation to cardiac electrical activity in epidemiological studies. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-12-18T23:16:46.75301-05:0
      DOI: 10.1002/jat.3092
  • HepaRG culture in tethered spheroids as an in vitro
           three‐dimensional model for drug safety screening
    • Authors: Zenan Wang; Xiaobei Luo, Chukwuemeka Anene‐Nzelu, Yu Yu, Xin Hong, Nisha Hari Singh, Lei Xia, Side Liu, Hanry Yu
      Abstract: Conventional two‐dimensional (2D) monolayer cultures of HepaRG cells allow in vitro maintenance of many liver‐specific functions. However, cellular dedifferentiation and functional deterioration over an extended culture period in the conventional 2D HepaRG culture have hampered its applications in drug testing. To address this issue, we developed tethered spheroids of HepaRG cells on Arg–Gly–Asp (RGD) and galactose‐conjugated substratum with an optimized hybrid ratio as an in vitro three‐dimensional (3D) human hepatocyte model. The liver‐specific gene expression level and drug metabolizing enzyme activities in HepaRG‐tethered spheorids were markedly higher than those in 2D cultures throughout the culture period of 7 days. The inducibility of three major cytochrome P450 (CYP) enzymes, namely CYP1A2, CYP2B6 and CYP3A4, was improved in both mRNA and activity level in tethered spheroids. Drug‐induced cytotoxic responses to model hepatotoxins (acetaminophen, chlorpromazine and ketoconazole) in tethered spheroids were comparable to 2D cultures as well as other studies in the literature. Our results suggested that the HepaRG‐tethered spheroid would be an alternative in vitro model suitable for drug safety screening. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-12-15T21:06:18.218232-05:
      DOI: 10.1002/jat.3090
  • Induction of the estrogen‐responsive genes encoding choriogenin H
           and L in the liver of male medaka (Oryzias latipes) upon exposure to
           estrogen receptor subtype‐selective ligands
    • Authors: Akemi Yamaguchi; Keisuke Kato, Koji Arizono, Nobuaki Tominaga
      Pages: n/a - n/a
      Abstract: Choriogenin (Chg) H and L are estrogen‐induced chorion precursors. We measured the induction of ChgH and ChgL mRNA in the livers of male medaka fish treated with Orthoester‐2k, a selective ligand for estrogen receptor (ER) α, and 2‐(4‐hydroxyphenyl)‐5‐hydroxy‐1,3‐benzoxazole (HPHB), a selective ligand of ERβ. Although both ChgH and ChgL mRNA were induced by treatment with Orthoester‐2k or HPHB separately, their combination induced much greater expression of each Chg. ChgH expression correlated more closely with Orthoester‐2k dosage when combined with a small fixed dose of HPHB (1 μm), whereas ChgL mRNA expression was more responsive to HPHB dose when combined with a fixed dose of Orthoester‐2k (2.8 nm). Moreover, upon long‐term treatment with Orthoester‐2k, ChgH mRNA and ERα mRNA expression showed similar patterns with peak expression between days 6 and 10. These results imply that ERβ primarily regulates ChgL mRNA expression and ERα action primarily regulates ChgH mRNA expression. Thus, it is necessary to develop screening methods for fish ER subtype‐specific ligands. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-12-10T08:14:40.91655-05:0
      DOI: 10.1002/jat.3063
  • Maternal exposure to 3,3’‐iminodipropionitrile targets
           late‐stage differentiation of hippocampal granule cell lineages to
           affect brain‐derived neurotrophic factor signaling and interneuron
           subpopulations in rat offspring
    • Authors: Megu Itahashi; Hajime Abe, Takeshi Tanaka, Sayaka Mizukami, Yoh Kikuchihara, Toshinori Yoshida, Makoto Shibutani
      Abstract: 3,3’‐Iminodipropionitrile (IDPN) causes neurofilament (NF)‐filled swellings in the proximal segments of many large‐caliber myelinated axons. This study investigated the effect of maternal exposure to IDPN on hippocampal neurogenesis in rat offspring using pregnant rats supplemented with 0 (controls), 67 or 200 ppm IDPN in drinking water from gestational day 6 to day 21 after delivery. On postnatal day (PND) 21, female offspring subjected to analysis had decreased parvalbumin+, reelin+ and phospho‐TrkB+ interneurons in the dentate hilus at 200 ppm and increased granule cell populations expressing immediate‐early gene products, Arc or c‐Fos, at ≥  67 ppm. mRNA expression in the dentate gyrus examined at 200 ppm decreased with brain‐derived neurotrophic factor (Bdnf) and very low density lipoprotein receptor. Immunoreactivity for phosphorylated NF heavy polypeptide decreased in the molecular layer of the dentate gyrus and the stratum radiatum of the cornu ammonis (CA) 3, portions showing axonal projections from mossy cells and pyramidal neurons, at 200 ppm on PND 21, whereas immunoreactivity for synaptophysin was unchanged in the dentate gyrus. Observed changes all disappeared on PND 77. There were no fluctuations in the numbers of apoptotic cells, proliferating cells and subpopulations of granule cell lineage in the subgranular zone on PND 21 and PND 77. Thus, maternal IDPN exposure may reversibly affect late‐stage differentiation of granule cell lineages involving neuronal plasticity as evident by immediate‐early gene responses to cause BDNF downregulation resulting in a reduction in parvalbumin+ or reelin+ interneurons and suppression of axonal plasticity in the mossy cells and CA3 pyramidal neurons. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-11-25T07:53:45.351994-05:
      DOI: 10.1002/jat.3086
  • Successful validation of genomic biomarkers for human immunotoxicity in
           Jurkat T cells in vitro
    • Authors: Peter C. J. Schmeits; Jia Shao, Danique A. Krieken, Oscar L. Volger, Henk Loveren, Ad. A. C. M. Peijnenburg, Peter J. M. Hendriksen
      Abstract: Previously, we identified 25 classifier genes that were able to assess immunotoxicity using human Jurkat T cells. The present study aimed to validate these classifiers. For that purpose, Jurkat cells were exposed for 6 h to subcytotoxic doses of nine immunotoxicants, five non‐immunotoxicants and four compounds for which human immunotoxicity has not yet been fully established. RNA was isolated and subjected to Fluidigm quantitative real time (qRT)–PCR analysis. The sensitivity, specificity and accuracy of the screening assay as based on the nine immunotoxicants and five non‐immunotoxicants used in this study were 100%, 80% and 93%, respectively, which is better than the performance in our previous study. Only one compound was classified as false positive (benzo‐e‐pyrene). Of the four potential (non‐)immunotoxicants, chlorantraniliprole and Hidrasec were classified immunotoxic and Sunset yellow and imidacloprid as non‐immunotoxic. ToxPi analysis of the PCR data provided insight in the molecular pathways that were affected by the compounds. The immunotoxicants 2,3‐dichloro‐propanol and cypermethrin, although structurally different, affected protein metabolism and cholesterol biosynthesis and transport. In addition, four compounds, i.e. chlorpyrifos, aldicarb, benzo‐e‐pyrene and anti‐CD3, affected genes in cholesterol metabolism and transport, protein metabolism and transcription regulation. qRT–PCR on eight additional genes coding for similar processes as defined in ToxPi analyzes, supported these results. In conclusion, the 25 immunotoxic classifiers performed very well in a screening with new non‐immunotoxic and immunotoxic compounds. Therefore, the Jurkat screening assay has great promise to be applied within a tiered approach for animal free testing of human immunotoxicity. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-11-25T07:38:41.511308-05:
      DOI: 10.1002/jat.3079
  • Surface‐expressed insulin receptors as well as IGF‐I receptors
           both contribute to the mitogenic effects of human insulin and its
    • Authors: Anders Lundby; Pernille Bolvig, Anne Charlotte Hegelund, Bo F. Hansen, Jesper Worm, Anne Lützen, Nils Billestrup, Christine Bonnesen, Martin B. Oleksiewicz
      Abstract: There is a medical need for new insulin analogues. Yet, molecular alterations to the insulin molecule can theoretically result in analogues with carcinogenic effects. Preclinical carcinogenicity risk assessment for insulin analogues rests to a large extent on mitogenicity assays in cell lines. We therefore optimized mitogenicity assay conditions for a panel of five cell lines. All cell lines expressed insulin receptors (IR), IGF‐I receptors (IGF‐IR) and hybrid receptors, and in all cell lines, insulin as well as the comparator compounds X10 and IGF‐I caused phosphorylation of the IR as well as IGF‐IR. Insulin exhibited mitogenicity EC50 values in the single‐digit nanomolar to picomolar range. We observed correlations across cell types between (i) mitogenic potency of insulin and IGF‐IR/IR ratio, (ii) Akt phosphorylation and mitogenic potency and (iii) Akt phosphorylation and IR phosphorylation. Using siRNA‐mediated knockdown of IR and IGF‐IR, we observed that in HCT 116 cells the IR appeared dominant in driving the mitogenic response to insulin, whereas in MCF7 cells the IGF‐IR appeared dominant in driving the mitogenic response to insulin. Together, our results show that the IR as well as IGF‐IR may contribute to the mitogenic potency of insulin. While insulin was a more potent mitogen than IGF‐I in cells expressing more IR than IGF‐IR, the hyper‐mitogenic insulin analogue X10 was a more potent mitogen than insulin across all cell types, supporting that the hyper‐mitogenic effect of X10 involves the IR as well as the IGF‐IR. These results are relevant for preclinical safety assessment of developmental insulin analogues. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-11-21T03:01:09.888264-05:
      DOI: 10.1002/jat.3082
  • Tributyltin alters secretion of interleukin 1 beta from human immune cells
    • Authors: Shyretha Brown; Margaret Whalen
      Abstract: Tributyltin (TBT) has been used as a biocide in industrial applications such as wood preservation, antifouling paint and antifungal agents. Owing to its many uses, it contaminates the environment and has been found in human blood samples. Interleukin‐1 beta (IL‐1β) is a pro‐inflammatory cytokine that promotes cell growth, tissue repair and immune response regulation. Produced predominately by both monocytes and macrophages, IL‐1β appears to increase the invasiveness of certain tumors. This study shows that TBT modifies the secretion of IL‐1β from increasingly reconstituted preparations of human immune cells. IL‐1β secretion was examined after 24‐, 48‐h or 6‐day exposures to TBT in highly enriched human natural killer (NK) cells, monocyte‐depleted peripheral blood mononuclear cells (MD‐PBMCs), PBMCs, granulocytes and a preparation combining both PBMCs and granulocytes (PBMCs+granulocytes). TBT altered IL‐1β secretion from all of the cell preparations. The 200 nM concentration of TBT normally blocked the secretion of IL‐1β, whereas lower concentrations (usually 5–50 nM) elevated secretion of IL‐1β. Examination of the signaling pathway(s) responsible for the elevated secretion of IL‐1β was carried out in MD‐PBMCs. Pathways examined were IL‐1β processing (Caspase‐1), mitogen‐activated protein kinases (MAPKs) and nuclear factor kappa B (NFκB). Results indicated that MAPK pathways (p44/42 and p38) appear to be the targets of TBT that lead to increased IL‐1β secretion from immune cells. These results from human immune cells show IL‐1β dysregulation by TBT is occurring ex vivo. Thus, the potential for in vivo effects on pro‐inflammatory cytokine levels may possibly be a consequence of TBT exposures. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-11-07T22:41:26.682232-05:
      DOI: 10.1002/jat.3087
  • 2, 3, 7, 8‐Tetrachlorodibenzo‐p‐dioxin induces premature
           senescence of astrocytes via WNT/β‐catenin signaling and ROS
    • Authors: Xiaoke Nie; Lingwei Liang, Hanqing Xi, Shengyang Jiang, Junkang Jiang, Cuiying Tang, Xipeng Liu, Suyi Liu, Chunhua Wan, Jianya Zhao, Jianbin Yang
      Abstract: 2, 3, 7, 8‐tetrachlorodibenzo‐p‐dioxin (TCDD) is a ubiquitous environmental contaminant that could exert significant neurotoxicity in the human nervous system. Nevertheless, the molecular mechanism underlying TCDD‐mediated neurotoxicity has not been clarified clearly. Herein, we investigated the potential role of TCDD in facilitating premature senescence in astrocytes and the underlying molecular mechanisms. Using the senescence‐associated β‐galactosidase (SA‐β‐Gal) assay, we demonstrated that TCDD exposure triggered significant premature senescence of astrocyte cells, which was accompanied by a marked activation of the Wingless and int (WNT)/β‐catenin signaling pathway. In addition, TCDD altered the expression of senescence marker proteins, such as p16, p21 and GFAP, which together have been reported to be upregulated in aging astrocytes, in both dose‐ and time‐dependent manners. Further, TCDD led to cell‐cycle arrest, F‐actin reorganization and the accumulation of cellular reactive oxygen species (ROS). Moreover, the ROS scavenger N‐acetylcysteine (NAC) markedly attenuated TCDD‐induced ROS production, cellular oxidative damage and astrocyte senescence. Notably, the application of XAV939, an inhibitor of WNT/β‐catenin signaling pathway, ameliorated the effect of TCDD on cellular β‐catenin level, ROS production, cellular oxidative damage and premature senescence in astrocytes. In summary, our findings indicated that TCDD might induce astrocyte senescence via WNT/β‐catenin and ROS‐dependent mechanisms. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-11-07T22:08:59.991905-05:
      DOI: 10.1002/jat.3084
  • Acute toxicity of 50 metals to Daphnia magna
    • Authors: Akira Okamoto; Masumi Yamamuro, Norihisa Tatarazako
      Abstract: Metals are essential for human life and physiological functions but may sometimes cause disorders. Therefore, we conducted acute toxicity testing of 50 metals in Daphnia magna: EC50s of seven elements (Be, Cu, Ag, Cd, Os, Au and Hg) were  100,000 µg l−1; and. 7 elements (Ti, Zr, Bi, Nb, Hf, Re and Ta) did not show EC50 at the upper limit of respective aqueous solubility, and EC50s were not obtained. Ga, Ru and Pd adhered to the body of D. magna and physically retarded the movement of D. magna. These metals formed hydroxides after adjusting the pH. Therefore, here, we distinguished this physical effect from the physiological toxic effect. The acute toxicity results of 40 elements obtained in this study were not correlated with electronegativity. Similarly, the acute toxicity results of metals including the rare metals were also not correlated with first ionization energy, atomic weight, atomic number, covalent radius, atomic radius or ionic radius. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-11-07T21:25:55.880885-05:
      DOI: 10.1002/jat.3078
  • RNA‐Seq‐based toxicogenomic assessment of fresh frozen and
           formalin‐fixed tissues yields similar mechanistic insights
    • Authors: Scott S. Auerbach; Dhiral P. Phadke, Deepak Mav, Stephanie Holmgren, Yuan Gao, Bin Xie, Joo Heon Shin, Ruchir R. Shah, B. Alex Merrick, Raymond R. Tice
      Abstract: Formalin‐fixed, paraffin‐embedded (FFPE) pathology specimens represent a potentially vast resource for transcriptomic‐based biomarker discovery. We present here a comparison of results from a whole transcriptome RNA‐Seq analysis of RNA extracted from fresh frozen and FFPE livers. The samples were derived from rats exposed to aflatoxin B1 (AFB1) and a corresponding set of control animals. Principal components analysis indicated that samples were separated in the two groups representing presence or absence of chemical exposure, both in fresh frozen and FFPE sample types. Sixty‐five percent of the differentially expressed transcripts (AFB1 vs. controls) in fresh frozen samples were also differentially expressed in FFPE samples (overlap significance: P 
      PubDate: 2014-11-06T04:16:58.370692-05:
      DOI: 10.1002/jat.3068
  • Impact of di‐ethylhexylphthalate exposure on metabolic programming
           in P19 ECC‐derived cardiomyocytes
    • Authors: Kristina Schaedlich; Juliane‐Susanne Schmidt, Wing Yee Kwong, Kevin D. Sinclair, Randy Kurz, Heinz‐Georg Jahnke, Bernd Fischer
      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.
      PubDate: 2014-10-29T01:04:56.648549-05:
      DOI: 10.1002/jat.3085
  • Impacts of the feed contaminant deoxynivalenol on the intestine of
           monogastric animals: poultry and swine
    • Authors: Khaled Ghareeb; Wageha A. Awad, Josef Böhm, Qendrim Zebeli
      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.
      PubDate: 2014-10-28T19:58:02.385342-05:
      DOI: 10.1002/jat.3083
  • Cellular uptake and toxicity effects of silver nanoparticles in mammalian
           kidney cells
    • Authors: Mirta Milić; Gerd Leitinger, Ivan Pavičić, Maja Zebić Avdičević, Slaven Dobrović, Walter Goessler, Ivana Vinković Vrček
      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.
      PubDate: 2014-10-28T19:21:35.91969-05:0
      DOI: 10.1002/jat.3081
  • Long‐term exposures to di‐n‐butyl phthalate inhibit body
           growth and impair gonad development in juvenile Murray rainbowfish
           (Melanotaenia fluviatilis)
    • Authors: Harpreet Bhatia; Anupama Kumar, John C. Chapman, Mike J. McLaughlin
      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.
      PubDate: 2014-10-28T01:12:32.140042-05:
      DOI: 10.1002/jat.3076
  • Organ‐specific distribution of gold nanoparticles by their surface
    • Authors: Jong Kwon Lee; Tae Sung Kim, Ji Young Bae, A. Young Jung, Sang Min Lee, Ji Hyun Seok, Hang Sik Roh, Chi Won Song, Mi Jin Choi, Jinyoung Jeong, Bong Hyun Chung, Yun‐Geon Lee, Jayoung Jeong, Wan‐Seob Cho
      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.
      PubDate: 2014-10-28T00:35:02.572138-05:
      DOI: 10.1002/jat.3075
  • Assessment of temperature‐induced hERG channel blockade variation by
    • Authors: Rahul R. Kauthale; Shruta S. Dadarkar, Raghib Husain, Vikas V. Karande, Madhumanjiri M. Gatne
      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.
      PubDate: 2014-10-28T00:14:26.431173-05:
      DOI: 10.1002/jat.3074
  • d‐α‐tocopheryl polyethylene glycol 1000
           succinate‐containing vehicles provide no detectable chemoprotection
           from oxidative damage
    • Authors: Bethany R. Baumgart; Terry R. Van Vleet, Damir Simic, Theodora W. Salcedo, Kimberley Lentz, Michael Donegan, Marc H. Davies, Roderick T. Bunch, Thomas P. Sanderson, Robert W. Lange
      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.
      PubDate: 2014-10-27T23:54:17.450279-05:
      DOI: 10.1002/jat.3072
  • Comparison of the kinetics of various biomarkers of benzo[a]pyrene
           exposure following different routes of entry in rats
    • Authors: Marjory Moreau; Michèle Bouchard
      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.
      PubDate: 2014-10-27T20:01:41.409244-05:
      DOI: 10.1002/jat.3070
  • 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
  • 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
    • 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
  • 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
  • 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
  • Sertoli cell as a model in male reproductive toxicology: Advantages and
    • Abstract: Pressure towards population aging in the demographic pyramid is not only due to sociological/personal choices but also due to subfertility or infertility. There are several chemicals and mixtures that impair male fertility. While experimental animal models are crucial to identify compounds that affect male fertility, it is essential to use reliable in vitro models to determine cellular targets and intracellular pathways that mediate chemical toxicity in the male reproductive system. In this review, we focused on the somatic Sertoli cell (SC) that, within the testis, is a major target for hormonal signaling and provides physical and nutritional support to developing germ cells. The different outcomes possible in each type of study: in vivo versus in vitro (either in primary or immortalized cell cultures) are analyzed. Herein, we intend to clarify the unique features that render SCs as excellent candidates for a robust in vitro model to study the deleterious effects of chemicals on male reproductive health. The sensitivity of SCs to toxicants/pharmaceuticals is discussed and, based on the literature reviewed we propose the in vitro study of SC physiology as a model to disclose deleterious effects of substances to male fertility. Copyright © 2015 John Wiley & Sons, Ltd.
  • Inhibitory effect of apocynin on methylglyoxal‐mediated glycation in
           osteoblastic MC3T3‐E1 cells
    • 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.
  • Synergistic cytotoxicity and DNA strand breaks in cells and plasmid DNA
           exposed to uranyl acetate and ultraviolet radiation
    • 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.
  • Carnosic acid induces autophagic cell death through inhibition of the
           Akt/mTOR pathway in human hepatoma cells
    • 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.
  • Heterozygous p53 knockout mouse model for dehydropyrrolizidine
           alkaloid‐induced carcinogenesis
    • Abstract: Dehydropyrrolizidine alkaloids (DHPA) are a large, structurally diverse group of plant‐derived protoxins that are potentially carcinogenic. With worldwide significance, these alkaloids can contaminate or be naturally present in the human food supply. To develop a small animal model that may be used to compare the carcinogenic potential of the various DHPAs, male heterozygous p53 knockout mice were administered a short‐term treatment of riddelliine 5, 15 or 45 mg kg–1 bodyweight day–1 by oral gavage for 14 days, or dosed a long‐term treatment of riddelliine 1 mg kg–1 bodyweight day–1 in pelleted feed for 12 months. Exposure to riddelliine increased the odds of tumor development in a dose‐responsive manner (odds ratio 2.05 and Wald 95% confidence limits between 1.2 and 3.4). The most common neoplastic process was hepatic hemangiosarcoma, which is consistent with published lifetime rodent riddelliine carcinogenesis studies. Angiectasis (peliosis hepatis) and other previously unreported lesions were also identified. The results of this research demonstrate the utility of the heterozygous p53 knockout mouse model for further investigation of comparative carcinogenesis of structurally and toxicologically different DHPAs and their N‐oxides. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.
  • The effect of a methyl‐deficient diet on the global DNA methylation
           and the DNA methylation regulatory pathways
    • Abstract: Methyl‐deficient diets are known to induce various liver disorders, in which DNA methylation changes are implicated. Recent studies have clarified the existence of the active DNA demethylation pathways that start with oxidization of 5‐methylcytosine (5meC) to 5‐hydroxymethylcytosine by ten‐eleven translocation (Tet) enzymes, followed by the action of base–excision–repair pathways. Here, we investigated the effects of a methionine–choline‐deficient (MCD) diet on the hepatic DNA methylation of mice by precisely quantifying 5meC using a liquid chromatography–electrospray ionization–mass spectrometry and by investigating the regulatory pathways, including DNA demethylation. Although feeding the MCD diet for 1 week induced hepatic steatosis and lower level of the methyl donor S‐adenosylmethionine, it did not cause a significant reduction in the 5meC content. On the other hand, the MCD diet significantly upregulated the gene expression of the Tet enzymes, Tet2 and Tet3, and the base–excision–repair enzymes, thymine DNA glycosylase and apurinic/apyrimidinic‐endonuclease 1. At the same time, the gene expression of DNA methyltransferase 1 and a, was also significantly increased by the MCD diet. These results suggest that the DNA methylation level is precisely regulated even when dietary methyl donors are restricted. Methyl‐deficient diets are well known to induce oxidative stress and the oxidative‐stress‐induced DNA damage, 8‐hydroxy‐2′‐deoxyguanosine (8OHdG), is reported to inhibit DNA methylation. In this study, we also clarified that the increase in 8OHdG number per DNA by the MCD diet is approximately 10 000 times smaller than the reduction in 5meC number, suggesting the contribution of 8OHdG formation to DNA methylation would not be significant. Copyright © 2015 John Wiley & Sons, Ltd.
  • Analysis of drugs of abuse in human plasma by dispersive
           liquid–liquid microextraction and high‐performance liquid
    • 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.
  • Cytotoxic and apoptotic activities of the plancitoxin I from the venom of
           crown‐of‐thorns starfish (Acanthaster planci) on A375.S2 cells
    • 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.
  • Systemic drugs inducing non‐immediate cutaneous adverse reactions
           and contact sensitizers evoke similar responses in THP‐1 cells
    • 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.
  • Elevated levels of antibodies against xenobiotics in a subgroup of healthy
    • 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.
  • Toxicity of new emerging pollutant tris‐(2,3‐dibromopropyl)
           isocyanurate on BALB/c mice
    • 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.
  • In utero and early childhood exposure to arsenic decreases lung function
           in children
    • 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.
  • Involvement of mitogen‐activated protein kinase and NF‐κB
           signaling pathways in perfluorooctane sulfonic acid‐induced
           inflammatory reaction in BV2 microglial cells
    • Abstract: Microglial activation is closely related to the pathogenesis of neurodegenerative diseases by producing proinflammatory cytokines. Perfluorooctane sulfonic acid (PFOS), known as an emerging persistent organic pollutant, is reported to disturb human immune homeostasis; however, whether it affects cytokine production or the immune response in the central nervous system remains unclear. The present study was aimed to explore whether PFOS contributed to inflammatory action and to investigate the corresponding mechanisms in BV2 microglia. PFOS‐mediated morphologic changes, cytokine responses and signaling events were examined by light microscopy, real‐time polymerase chain reaction, enzyme‐linked immunosorbent assay and Western blot assays. Our results indicated that PFOS increased BV2 cells activation and simultaneously increased tumor necrosis factor alpha and interleukin‐6 expression. In addition, the c‐Jun N‐terminal protein kinase inhibitor (SP600125), as well as ERK1/2 blocker (PD98059), transcriptionally at least, displayed anti‐inflammatory properties on PFOS‐elicited cytokine responses. Moreover, the inflammatory transcription factor NF‐κB was specifically activated by PFOS as well. These results, taken together, suggested that PFOS exerts its functional effects on the response of microglial cell activation via, in part, the c‐Jun N‐terminal protein kinase, ERK and NF‐κB signaling pathways with its subsequent influence on proinflammatory action. Copyright © 2015 John Wiley & Sons, Ltd.
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