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  Subjects -> ENVIRONMENTAL STUDIES (Total: 813 journals)
    - ENVIRONMENTAL STUDIES (740 journals)
    - POLLUTION (22 journals)
    - WASTE MANAGEMENT (10 journals)

ENVIRONMENTAL STUDIES (740 journals)            First | 1 2 3 4 5 6 7 8     

Hydrology: Current Research     Open Access   (Followers: 11)
IAMURE International Journal of Ecology and Conservation     Open Access   (Followers: 3)
Ideas in Ecology and Evolution     Open Access   (Followers: 10)
IEEE Transactions on Network and Service Management     Hybrid Journal   (Followers: 10)
IMA Journal of Management Mathematics     Hybrid Journal   (Followers: 1)
Indiana Journal of Global Legal Studies     Full-text available via subscription   (Followers: 2)
Indoor Air     Hybrid Journal   (Followers: 1)
Information Systems Management     Hybrid Journal   (Followers: 17)
Information Technology and Management     Hybrid Journal   (Followers: 9)
IngenierĂ­a HidrĂ¡ulica y Ambiental     Open Access  
Inhalation Toxicology     Hybrid Journal   (Followers: 7)
Integrated Environmental Assessment and Management     Hybrid Journal   (Followers: 5)
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: 1)
International Journal of African Renaissance Studies - Multi-, Inter- and Transdisciplinarity     Hybrid Journal   (Followers: 2)
International Journal of Agricultural and Environmental Information Systems     Full-text available via subscription   (Followers: 1)
International Journal of Alternative Propulsion     Hybrid Journal   (Followers: 1)
International Journal of Applied Psychoanalytic Studies     Hybrid Journal   (Followers: 2)
International Journal of Chinese Culture and Management     Hybrid Journal   (Followers: 1)
International Journal of Corrosion     Open Access   (Followers: 11)
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  
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: 7)
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: 11)
International Journal of Environmental Protection     Open Access   (Followers: 14)
International Journal of Environmental Research and Public Health     Open Access   (Followers: 16)
International Journal of Environmental Science and Technology     Hybrid Journal   (Followers: 6)
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: 4)
International Journal of Global Environmental Issues     Hybrid Journal   (Followers: 4)
International Journal of Global Warming     Hybrid Journal   (Followers: 5)
International Journal of Greenhouse Gas Control     Partially Free   (Followers: 6)
International Journal of Health Planning and Management     Hybrid Journal   (Followers: 6)
International Journal of Hygiene and Environmental Health     Hybrid Journal   (Followers: 6)
International Journal of Logistics Research and Applications : A Leading Journal of Supply Chain Management     Hybrid Journal   (Followers: 10)
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: 2)
International Journal of Regulation and Governance     Hybrid Journal   (Followers: 2)
International Journal of Reliability and Safety     Hybrid Journal   (Followers: 7)
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: 10)
International Journal of Sustainable Construction Engineering and Technology     Open Access   (Followers: 10)
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: 11)
Interventions : International Journal of Postcolonial Studies     Hybrid Journal   (Followers: 10)
IOP Conference Series: Earth and Environmental Science     Open Access   (Followers: 7)
Iranian Studies     Hybrid Journal   (Followers: 10)
Irish Educational Studies     Hybrid Journal   (Followers: 2)
Irish Journal of Earth Sciences     Full-text available via subscription  
Irish Political Studies     Hybrid Journal   (Followers: 10)
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)
Italian Studies     Hybrid Journal   (Followers: 6)
Jahangirnagar University Environmental Bulletin     Open Access  
Journal of Bioremediation & Biodegradation     Open Access   (Followers: 2)
Journal of Earth Science & Climatic Change     Open Access   (Followers: 7)
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   (Followers: 1)
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   (Followers: 1)
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: 133)

  First | 1 2 3 4 5 6 7 8     

Journal Cover   Journal of Applied Toxicology
  [SJR: 0.799]   [H-I: 53]   [10 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  [1607 journals]
  • Toxicogenomic responses of human liver HepG2 cells to silver nanoparticles
    • Authors: Saura C. Sahu; Jiwen Zheng, Jeffrey J. Yourick, Robert L. Sprando, Xiugong Gao
      Abstract: The increased use of silver nanoparticles (AgNPs) in foods and cosmetics has raised public safety concerns. However, only limited knowledge exists on the effect of AgNPs on the cellular transcriptome. This study evaluated global gene expression profiles of human liver HepG2 cells exposed to 20 and 50 nm AgNPs for 4 and 24 h at 2.5 µg ml–1. Exposure to 20 nm AgNPs resulted in 811 altered genes after 4 h, but much less after 24 h. Exposure to 50 nm AgNPs showed minimal altered genes at both exposure times. The HepG2 cells responded to the toxic insult of AgNPs by transiently upregulating stress response genes such as metallothioneins and heat shock proteins. Functional analysis of the altered genes showed more than 20 major biological processes were affected, of which metabolism, development, cell differentiation and cell death were the most dominant categories. Several cellular pathways were also impacted by AgNP exposure, including the p53 signaling pathway and the NRF2‐mediated oxidative stress response pathway, which may lead to increased oxidative stress and DNA damage in the cell and potentially result in genotoxicity and carcinogenicity. Together, these results indicate that HepG2 cells underwent a multitude of cellular processes in response to the toxic insult of AgNP exposure, and suggest that toxicogenomic characterization of human HepG2 cells could serve as an alternative model for assessing toxicities of NPs. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.
      PubDate: 2015-05-26T00:18:17.224014-05:
      DOI: 10.1002/jat.3170
  • Short‐term, low‐dose cadmium exposure induces
           hyperpermeability in human renal glomerular endothelial cells
    • Authors: Liqun Li; Fengyun Dong, Dongmei Xu, Linna Du, Suhua Yan, Hesheng Hu, Corrinne G. Lobe, Fan Yi, Carolyn M. Kapron, Ju Liu
      Abstract: The kidney is the principal organ targeted by exposure to cadmium (Cd), a well‐known toxic metal. Even at a low level, Cd damages glomerular filtration. However, little is known about the effects of Cd on the glomerular endothelium, which performs the filtration function and directly interacts with Cd in blood plasma. In this study, we cultured human renal glomerular endothelial cells (HRGECs) in the presence of serum with treatment of a short term (1 h) and low concentration (1 μm) of Cd, which mimics the pattern of glomerular endothelium exposure to Cd in vivo. We found that this short‐term, low‐dose Cd exposure does not induce cytotoxicity, but increases permeability in HRGECs monolayers and redistributes adherens junction proteins vascular endothelial‐cadherin and β‐catenin. Though short‐term, low‐dose Cd exposure activates all three major mitogen activated protein kinases, only the inhibitor of p38 mitogen activated protein kinase partially prevents Cd‐induced hyperpermeability in HRGECs. Our data indicate that the presence of Cd in blood circulation might directly disrupt the glomerular endothelial cell barrier and contribute to the development of clinical symptoms of glomerular diseases. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-25T21:57:08.646349-05:
      DOI: 10.1002/jat.3168
  • Onset of hepatocarcinogen‐specific cell proliferation and cell cycle
           aberration during the early stage of repeated hepatocarcinogen
           administration in rats
    • Authors: Masayuki Kimura; Hajime Abe, Sayaka Mizukami, Takeshi Tanaka, Megu Itahashi, Nobuhiko Onda, Toshinori Yoshida, Makoto Shibutani
      Abstract: We have previously reported that a 28‐day treatment of carcinogens evoking target cell proliferation activates G1/S checkpoint function and apoptosis, as well as induction of aberrant ubiquitin D (Ubd) expression, suggesting disruptive spindle checkpoint function, in rats. The present study aimed to determine the onset time of rat liver cells to undergo carcinogen‐specific cell cycle aberration and proliferation. Animals were treated orally with a hepatocarcinogenic dose of methyleugenol or thioacetamide for 3, 7 or 28 days. For comparison, some animals were subjected to partial hepatectomy or treated with noncarcinogenic hepatotoxicants (acetaminophen, α‐naphthyl isothiocyanate or promethazine). Carcinogen‐specific liver cell kinetics appeared at day 28 as evident by increases of cell proliferation, p21Cip1+ cells, phosphorylated‐Mdm2+ cells and cleaved caspase 3+ cells, and upregulation of DNA damage‐related genes. Hepatocarcinogens also downregulated Rbl2 and upregulated Cdkn1a and Mdm2, and decreased Ubd+ cells co‐expressing phosphorylated‐histone H3 (p‐Histone H3) and p‐Histone H3+ cell ratio within the Ki‐67+ proliferating population. These results suggest that it takes 28 days to induce hepatocarcinogen‐specific early withdrawal of proliferating cells from M phase due to disruptive spindle checkpoint function as evidenced by reduction of Ubd+ cells staying at M phase. Disruption of G1/S checkpoint function reflected by downregulation of Rbl2 as well as upregulation of Mdm2 suggestive of sequestration of retinoblastoma protein is responsible for the facilitation of carcinogen‐induced cell proliferation at day 28. Accumulation of DNA damage probably in association with facilitation of p53 degradation by activation of Mdm2 may be a prerequisite for aberrant p21Cip1 activation, which is responsible for apoptosis. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-22T07:41:41.042646-05:
      DOI: 10.1002/jat.3163
  • RNA sequencing provides insights into the toxicogenomic response of ZF4
           cells to methyl methanesulfonate
    • Authors: Zhouquan Li; Yong Long, Liqiao Zhong, Guili Song, Xiaohua Zhang, Li Yuan, Zongbin Cui, Heping Dai
      Abstract: Whole genome transcriptomic studies are powerful for characterizing the molecular mechanisms underlying the physiological effects of chemicals, and are informative for environmental health risk assessment. Alkylating agents are an abundant class of chemicals that can damage DNA in the environment, and are used for anticancer treatments. Currently, little is known regarding the molecular mechanisms of toxic alkylating agents in zebrafish cell lines. In this study, RNA‐sequencing was used to investigate the transcriptomic responses of zebrafish ZF4 cells following exposure to the model genotoxicant methyl methanesulfonate (MMS). The half‐maximal inhibitory concentration (IC50) of MMS was 639.16 ± 61.8 µm, and apoptosis was induced within 24 h of exposure. RNA sequencing identified 3601 differentially expressed genes (DEGs) that were upregulated and 3037 that were downregulated. Gene ontology enrichment analysis revealed that most DEGs belonged to synthesis and metabolism categories. RNA‐associated processes were the most upregulated, while cell cycle and adhesion were the most repressed processes, and neuron‐related processes were the most downregulated developmental process. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis identified DNA damage repair, cell cycle, apoptosis and spliceosome as overrepresented terms. Six types of alternative splicing were detected. In total, 1156 alternative splicing DEGs were specifically expressed following MMS treatment, many of which belonged to metabolism and catabolic process categories. Cluster analysis of orthologs was able to extrapolate toxicotranscriptomic data between zebrafish and yeast. These results provide insight into the genome‐wide response of ZF4 cells following exposure to MMS, and this knowledge will inform future toxicogenomic data analysis and environmental health risk assessment. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-22T07:22:45.239857-05:
      DOI: 10.1002/jat.3147
  • A method for estimating the glomerular filtration rate in conscious
    • Authors: Hiroshi Satoh; Nana Nomiya, Daiki Imai, Shigeru Sato, Ken Sakurai, Kiyoshi Takasuna, Kazuhisa Furuhama
      Abstract: To establish a method for estimating the glomerular filtration rate (GFR) in conscious monkeys, the radiographic contrast medium iodixanol and the standard agent inulin were coadministered as tracers to male cynomolgus monkeys (Macaca fascicularis) as a bolus injection; blood was collected after 60, 90 and 120 min. An equation based on a single‐blood‐sample method derived from Jacobsson's formula was prepared using the data from healthy and saline‐ and gentamicin‐treated monkeys by a multisample strategy with iodixanol. The GFR using the equation with iodixanol was in agreement with that from the multisample method with inulin or iodixanol. When the GFR decreased to more than 60% of the basal reference level, serum creatinine concentrations tended to increase, whereas serum blood urea nitrogen concentrations fluctuated. The results suggest that the single‐sample‐blood method with iodixanol is a practical tool for estimating the monkey GFR in a toxicological research setting therefore minimizing animal sufferings. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-21T00:40:24.316596-05:
      DOI: 10.1002/jat.3178
  • Developmental toxicity and endocrine disruption of naphthenic acids on the
           early life stage of zebrafish (Danio rerio)
    • Authors: Jie Wang; Xiaofeng Cao, Yi Huang, Xiaoyan Tang
      Abstract: Oil sands process‐affected water (OSPW) has been reported to exhibit adverse effects on the environment and wildlife. Although the compounds responsible are unknown, naphthenic acids (NAs) have been considered to be implicated. The current study was designed to investigate whether NAs might cause developmental toxicity and endocrine disruption on the early life stage of zebrafish (Danio rerio). The success of embryo hatch was inhibited by 2.5 mg l–1 oil sands NAs (OS‐NAs) exposure, and both OSPW NAs and commercial NAs (C‐NAs) exposure resulted in a variety of developmental lesions in the fish larvae, such as yolk sac edema, pericardial edema and spinal malformation. The transcription of genes involved cytochrome P450 aromatase (CYP19a and CYP19b), estrogen receptors (ERα, ERβ1 and ERβ2), and vitellogenin (VTG) was analyzed to evaluate the endocrine disrupting effects of NAs. Significant up‐regulated gene expressions of CYP19b, ERα and VTG were observed in both OS‐NAs and C‐NAs groups, which indicated the deleteriously estrogenic potential of NAs. These results confirmed that NAs derived from crude petroleum could negatively impact the development and endocrine function of zebrafish, and be primarily responsible for the toxicity of OSPW. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-21T00:36:44.988264-05:
      DOI: 10.1002/jat.3166
  • Directional and color preference in adult zebrafish: Implications in
           behavioral and learning assays in neurotoxicology studies
    • Authors: Zachary A. Bault; Samuel M. Peterson, Jennifer L. Freeman
      Abstract: The zebrafish (Danio rerio) is a useful vertebrate model organism for neurological studies. While a number of behavior and learning assays are recently reported in the literature for zebrafish, many of these assays are still being refined. The initial purpose of this study was to apply a published T‐maze assay for adult zebrafish that measures how quickly an organism can discriminate between different color stimuli after receiving reinforcement to measure learning in a study investigating the later life impacts of developmental Pb exposure. The original results were inconclusive as the control group showed a directional and color preference. To assess directional preference further, a three‐chambered testing apparatus was constructed and rotated in several directions. The directional preference observed in males was alleviated by rotating the arms pointing west and east. In addition, color preference was investigated using all combinations of five different colors (orange, yellow, green, blue and purple). With directional preference alleviated results showed that both male and female zebrafish preferred colors of shorter wavelengths. An additional experiment tested changes in color preference due to developmental exposure to Pb in adult male zebrafish. Results revealed that Pb‐exposed males gained and lost certain color preferences compared to control males and the preference for short wavelengths was decreased. Overall, these results show that consideration and pretesting should be completed before applying behavioral and learning assays involving adult zebrafish to avoid innate preferences and confounding changes in neurotoxicology studies and that developmental Pb exposure alters color preferences in adult male zebrafish. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-20T21:12:27.765722-05:
      DOI: 10.1002/jat.3169
  • Effect‐directed identification of endocrine disruptors in plastic
           baby teethers
    • Authors: Elisabeth Berger; Theodoros Potouridis, Astrid Haeger, Wilhelm Püttmann, Martin Wagner
      Abstract: Concerns have been raised regarding the human health effects of endocrine disrupting chemicals (EDCs), many of which are associated with and leaching from plastics. As infants are particularly vulnerable to EDCs, we have investigated whether plastic teethers for babies represent a relevant source of exposure. Applying effect‐directed analysis, we use bioassays to screen teethers, toys used to soothe a baby's teething ache, for endocrine activity and chemical analysis to identify the causative compounds. We detected significant endocrine activity in two of 10 plastic teethers. Those samples leached estrogenic and/or antiandrogenic activity as detected in the Yeast Estrogen Screen and Yeast Antiandrogen Screen. After sample fractionation, gas chromatography–mass spectrometry non‐target screening revealed that methyl‐, ethyl‐ and propylparaben were responsible for the observed estrogenic and antiandrogenic activity in one product. The second product is likely to contain at least six different antiandrogenic compounds that remain so far unidentified. This study demonstrates that plastic teethers can be a source of infant exposure to well‐established and unknown EDCs. Because of their limited value to the product, but potential toxicity, manufacturers should critically revisit the use of parabens in plastic teethers and further toys. Moreover, plastic teethers might leach EDCs that escape routine analysis and, thus, toxicological evaluation. The resulting uncertainty in product safety poses a problem to consumers, producers and regulators that remain to be resolved. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-18T07:27:48.753651-05:
      DOI: 10.1002/jat.3159
  • Acute and subchronic toxicity of 20  kHz and
           60  kHz magnetic fields in rats
    • Authors: Izumi Nishimura; Atsushi Oshima, Kazumoto Shibuya, Takashi Mitani, Tadashi Negishi
      Abstract: Despite increasing use of intermediate frequency (IF) magnetic fields (MFs) in occupational and domestic settings, scientific evidence necessary for health risk assessments of IF MF is insufficient. Male and female Crl:CD(SD) rats (12 per sex per group) were exposed to 20 kHz, 0.20 mT(root mean square, rms) or 60 kHz, 0.10 mT(rms) sinusoidal MFs for 22 h day−1 for 14 days (acute) or 13 weeks (subchronic). Experiments were duplicated for each frequency to ensure outcome reproducibility, and examinations were blinded for quality assurance. All rats survived without significant clinical signs until the end of experiments. Some changes in body weight between the MF‐exposed and control groups were observed over the course of exposure, although the directions of the changes were inconsistent and not statistically significant after subchronic exposure. There were significant differences between MF‐exposed and control groups in some organ weights and parameters in hematology and clinical chemistry, but these were minor in magnitude and not repeated in duplicate experiments. Histopathological findings reflecting toxicity were sporadic. Frequencies of other findings were similar to historic data in this rat strain, and findings had no specific relationship to changes in organ weight or parameters of hematology and clinical chemistry in each animal. The changes observed throughout this study were considered biologically isolated and were attributable to chance associations rather than to MF exposure. The results, in particular the histopathological evidence, indicate an absence of toxicity in IF MF‐exposed rats and do not support the hypothesis that IF MF exposure produces significant toxicity. Copyright © 2015. The
      Authors . Journal of Applied Toxicology Published by John Wiley & Sons Ltd.
      PubDate: 2015-05-17T23:57:37.168612-05:
      DOI: 10.1002/jat.3161
  • Silver nanoparticles affect the neural development of zebrafish embryos
    • Authors: Qi Xin; Jeanette M. Rotchell, Jinping Cheng, Jun Yi, Qiang Zhang
      Abstract: Silver nanoparticles (AgNPs) have been widely used in commercial products. This study aims to understand the impact of AgNPs on the early developmental stages in zebrafish (Danio rerio) embryos. Embryos were exposed to two sizes of AgNPs at three dose levels, as well to free Ag+ ions, for a range of 4–96 h post‐fertilization (hpf). The acute exposure study showed that exposure to AgNPs affected the neurological development, and the exposed embryos exhibited anomalies such as small head with hypoplastic hindbrain, small eye and cardiac defects. At the molecular level, AgNPs altered the expression profiles of neural development‐related genes (gfap, huC and ngn1), metal‐sensitive metallothioneins and ABCC genes in exposed embryos. The expression of AhR2 and Cyp1A, which are usually considered to mediate polycyclic aromatic hydrocarbon toxicity, were also significantly changed. A size‐dependent uptake of AgNPs was observed, whereby 4 nm AgNPs were more efficiently taken up compared with the 10 nm‐sized particles. Importantly, the head area accumulated AgNPs more efficiently than the trunk area of exposed zebrafish embryos. No free Ag+ ions, which can be potentially released from the AgNP solutions, were detected. This study suggests that AgNPs could affect the neural development of zebrafish embryos, and the toxicity of AgNPs may be partially attributed to the comparatively higher uptake in the head area. These results indicate the potential neurotoxicity of AgNPs and could be extended to other aquatic organisms. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-15T00:29:17.852587-05:
      DOI: 10.1002/jat.3164
  • Intracellular calcium levels as screening tool for nanoparticle toxicity
    • Authors: Claudia Meindl; Tatjana Kueznik, Martina Bösch, Eva Roblegg, Eleonore Fröhlich
      Abstract: The use of engineered nano‐sized materials led to revolutionary developments in many industrial applications and in the medical field. These materials, however, also may cause cytotoxicity. In addition to size, surface properties and shape were identified as relevant parameters for cell damage. Cell damage may occur as disruption of membrane integrity, induction of apoptosis and by organelle damage. Generation of oxidative stress may serve as an indicator for cytotoxicity. Effects occurring upon short contact of particles with cells, for instance in the systemic blood circulation, could be identified according to increases of intracellular [Ca2+] levels, which are caused by variety of toxic stimuli. Negatively charged, neutral and positively charged polystyrene particles of different sizes were used to study the role of size and surface properties on viability, membrane disruption, apoptosis, lysosome function, intracellular [Ca2+] levels and generation of oxidative stress. Silica particles served to test this hypothesis. Twenty nm polystyrene particles as well as 12 nm and 40 nm silica particles caused membrane damage and apoptosis with no preference of the surface charge. Only 20 nm plain and amine functionalized polystyrene particles cause oxidative stress and only the plain particles lysosomal damage. A potential role of surface charge was identified for 200 nm polystyrene particles, where only the amidine particles caused lysosomal damage. Increases in intracellular [Ca2+] levels and cytotoxicity after 24 h was often linked but determination of intracellular [Ca2+] levels could serve to characterize further the type of membrane damage. © 2015 The
      Authors . Journal of Applied Toxicology Published by John Wiley & Sons Ltd.
      PubDate: 2015-05-14T21:12:31.32087-05:0
      DOI: 10.1002/jat.3160
  • Toxicological effects of pet food ingredients on canine bone
           marrow‐derived mesenchymal stem cells and enterocyte‐like
    • Authors: M. T. Ortega; B. Jeffery, J. E. Riviere, N. A. Monteiro‐Riviere
      Abstract: We developed an in vitro method to assess pet food ingredients safety. Canine bone marrow‐derived mesenchymal stem cells (BMSC) were differentiated into enterocyte‐like cells (ELC) to assess toxicity in cells representing similar patterns of exposure in vivo. The toxicological profile of clove leave oil, eugenol, guanosine monophosphate (GMP), GMP + inosine monophosphate, sorbose, ginger root extract, cinnamon bark oil, cinnamaldehyde, thyme oil, thymol and citric acid was assessed in BMSC and ELC. The LC50 for GMP + inosine monophosphate was 59.42 ± 0.90 and 56.7 ± 3.5 mg ml–1 for BMSC and ELC; 56.84 ± 0.95 and 53.66 ± 1.36 mg ml–1 for GMP; 0.02 ± 0.001 and 1.25 ± 0.47 mg ml–1 for citric acid; 0.077 ± 0.002 and 0.037 ± 0.01 mg ml–1 for cinnamaldehyde; 0.002 ± 0.0001 and 0.002 ± 0.0008 mg ml–1 for thymol; 0.080 ± 0.003 and 0.059 ± 0.001 mg ml–1 for thyme oil; 0.111 ± 0.002 and 0.054 ± 0.01 mg ml–1 for cinnamon bark oil; 0.119 ± 0.0004 and 0.099 ± 0.011 mg ml–1 for clove leave oil; 0.04 ± 0.001 and 0.028 ± 0.002 mg ml–1 for eugenol; 2.80 ± 0.11 and 1.75 ± 0.51 mg ml–1 for ginger root extract; > 200 and 116.78 ± 7.35 mg ml–1 for sorbose. Lemon grass oil was evaluated at 0.003–0.9 in BMSC and .03‐0.9 mg ml–1 in ELC and its mechanistic effect was investigated. The gene toxicology studies showed regulation of 61% genes in CYP450 pathway, 37% in cholestasis and 33% in immunotoxicity pathways for BMSC. For ELC, 80% for heat shock response, 69% for beta‐oxidation and 65% for mitochondrial energy metabolism. In conclusion, these studies provide a baseline against which differential toxicity of dietary feed ingredients can be assessed in vitro for direct effects on canine cells and demonstrate differential toxicity in differentiated cells that represent gastrointestinal epithelial cells. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-14T18:22:26.588619-05:
      DOI: 10.1002/jat.3158
  • Safety data on 19 vehicles for use in 1 month oral rodent
           pre‐clinical studies: administration of
           hydroxypropyl‐ß‐cyclodextrin causes renal toxicity
    • Authors: Guy Healing; Tabassum Sulemann, Peter Cotton, Jayne Harris, Adam Hargreaves, Rowena Finney, Sarah Kirk, Carolin Schramm, Clare Garner, Perrine Pivette, Lisa Burdett
      Abstract: Potential new drugs are assessed in pre‐clinical in vivo studies to determine their safety profiles. The drugs are formulated in vehicles suitable for the route of administration and the physicochemical properties of the drug, aiming to achieve optimal exposure in the test species. The availability of safety data on vehicles is often limited (incomplete data, access restricted/private databases). Nineteen potentially useful vehicles that contained new and/or increased concentrations of excipients and for which little safety data have been published were tested. Vehicles were dosed orally once daily to HanWistar rats for a minimum of 28 days and a wide range of toxicological parameters were assessed. Only 30% (w/v) hydroxypropyl‐ß‐cyclodextrin was found unsuitable owing to effects on liver enzymes (AST, ALT and GLDH), urinary volume and the kidneys (tubular vacuolation and tubular pigment). 20% (v/v) oleic acid caused increased salivation and hence this vehicle should be used with caution. As 40% (v/v) tetraethylene glycol affected urinary parameters, its use should be carefully considered, particularly for compounds suspected to impact the renal system and studies longer than 1 month. There were no toxicologically significant findings with 10% (v/v) dimethyl sulphoxide, 20% (v/v) propylene glycol, 33% (v/v) Miglyol®812, 20% (w/v) Kolliphor®RH40, 10% (w/v) Poloxamer 407, 5% (w/v) polyvinylpyrrolidone K30 or 10% (v/v) Labrafil®M1944. All other vehicles tested caused isolated or low magnitude effects which would not prevent their use. The aim of sharing these data, including adverse findings, is to provide meaningful information for vehicle selection, thereby avoiding repetition of animal experimentation. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-10T20:18:00.800754-05:
      DOI: 10.1002/jat.3155
  • Maternal exposure to hexachlorophene targets intermediate‐stage
           progenitor cells in the hippocampal neurogenesis involving myelin
           vacuolation of cholinergic and glutamatergic inputs in mice
    • Authors: Mizuho Kato; Hajime Abe, Megu Itahashi, Yoh Kikuchihara, Masayuki Kimura, Sayaka Mizukami, Toshinori Yoshida, Makoto Shibutani
      Abstract: Hexachlorophene (HCP) has been shown to induce myelin vacuolation due to intramyelinic edema of the nerve fibers in animal neural tissue. We investigated the maternal exposure effect of HCP on hippocampal neurogenesis in the offspring of pregnant mice supplemented with 0 (control), 33 or 100 ppm HCP in diet from gestational day 6 to day 21 after delivery. On postnatal day (PND) 21, offspring as examined in males exhibited decreased granule cell lineage populations expressing paired box 6, sex‐determining region Y‐box 2 and eomesodermin in the hippocampal subgranular zone (SGZ) accompanied by myelin vacuolation involving white matter tracts of the hippocampal fimbria at ≥ 33 ppm. However, SGZ cellular populations expressing brain lipid binding protein and doublecortin were unchanged at any dose. Transcript expression of cholinergic receptor genes, Chrna4 and Chrnb2, and glutamate receptor genes, Grm1 and Grin2d, examined at 100 ppm, decreased in the dentate gyrus. HCP exposure did not alter the number of proliferating or apoptotic cells in the SGZ, or reelin‐ or calcium‐binding protein‐expressing γ‐aminobutyric acid (GABA)ergic interneurons in the dentate hilus, on PND 21 and PND 77. All neurogenesis‐related changes observed in HCP‐exposed offspring on PND 21 disappeared on PND 77, suggesting that maternal HCP exposure at ≥ 33 ppm reversibly decreased type 2 intermediate‐stage progenitor cells in the hippocampal neurogenesis. Myelin vacuolation might be responsible for changes in neurogenesis possibly by reducing nerve conduction velocity of cholinergic inputs from the septal–hippocampal pathway to granule cell lineages and/or GABAergic interneurons, and of glutamatergic inputs to granule cell lineages. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-05-05T09:13:26.853194-05:
      DOI: 10.1002/jat.3162
  • Adverse Outcome Pathways can drive non‐animal approaches for safety
    • Authors: Natalie Burden; Fiona Sewell, Melvin E. Andersen, Alan Boobis, J. Kevin Chipman, Mark T. D. Cronin, Thomas H. Hutchinson, Ian Kimber, Maurice Whelan
      Abstract: Adverse Outcome Pathways (AOPs) provide an opportunity to develop new and more accurate safety assessment processes for drugs and other chemicals, and may ultimately play an important role in regulatory decision making. Not only can the development and application of AOPs pave the way for the development of improved evidence‐based approaches for hazard and risk assessment, there is also the promise of a significant impact on animal welfare, with a reduced reliance on animal‐based methods. The establishment of a useable and coherent knowledge framework under which AOPs will be developed and applied has been a first critical step towards realizing this opportunity. This article explores how the development of AOPs under this framework, and their application in practice, could benefit the science and practice of safety assessment, while in parallel stimulating a move away from traditional methods towards an increased acceptance of non‐animal approaches. We discuss here the key areas where current, and future initiatives should be focused to enable the translation of AOPs into routine chemical safety assessment, and lasting 3Rs benefits. © 2015 The
      Authors . Journal of Applied Toxicology published by John Wiley & Sons Ltd.
      PubDate: 2015-05-05T08:56:05.682065-05:
      DOI: 10.1002/jat.3165
  • mRNAs and miRNAs in whole blood associated with lung hyperplasia,
           fibrosis, and bronchiolo‐alveolar adenoma and adenocarcinoma after
           multi‐walled carbon nanotube inhalation exposure in mice
    • Authors: Brandi N. Snyder‐Talkington; Chunlin Dong, Linda M. Sargent, Dale W. Porter, Lauren M. Staska, Ann F. Hubbs, Rebecca Raese, Walter McKinney, Bean T. Chen, Lori Battelli, David T. Lowry, Steven H. Reynolds, Vincent Castranova, Yong Qian, Nancy L. Guo
      Abstract: Inhalation exposure to multi‐walled carbon nanotubes (MWCNT) in mice results in inflammation, fibrosis and the promotion of lung adenocarcinoma; however, the molecular basis behind these pathologies is unknown. This study determined global mRNA and miRNA profiles in whole blood from mice exposed by inhalation to MWCNT that correlated with the presence of lung hyperplasia, fibrosis, and bronchiolo‐alveolar adenoma and adenocarcinoma. Six‐week‐old, male, B6C3F1 mice received a single intraperitoneal injection of either the DNA‐damaging agent methylcholanthrene (MCA, 10 µg g−1 body weight) or vehicle (corn oil). One week after injections, mice were exposed by inhalation to MWCNT (5 mg m−3, 5 hours per day, 5 days per week) or filtered air (control) for a total of 15 days. At 17 months post‐exposure, mice were euthanized and examined for the development of pathological changes in the lung, and whole blood was collected and analyzed using microarray analysis for global mRNA and miRNA expression. Numerous mRNAs and miRNAs in the blood were significantly up‐ or down‐regulated in animals developing pathological changes in the lung after MCA/corn oil administration followed by MWCNT/air inhalation, including fcrl5 and miR‐122‐5p in the presence of hyperplasia, mthfd2 and miR‐206‐3p in the presence of fibrosis, fam178a and miR‐130a‐3p in the presence of bronchiolo‐alveolar adenoma, and il7r and miR‐210‐3p in the presence of bronchiolo‐alveolar adenocarcinoma, among others. The changes in miRNA and mRNA expression, and their respective regulatory networks, identified in this study may potentially serve as blood biomarkers for MWCNT‐induced lung pathological changes. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-29T23:36:08.828959-05:
      DOI: 10.1002/jat.3157
  • In vitro neurotoxicity evaluation of piperazine designer drugs in
           differentiated human neuroblastoma SH‐SY5Y cells
    • Authors: M. D. Arbo; R. Silva, D. J. Barbosa, D. Dias Silva, S. P. Silva, J. P. Teixeira, M. L. Bastos, H. Carmo
      Abstract: Abuse of synthetic drugs is widespread worldwide. Studies indicate that piperazine designer drugs act as substrates at dopaminergic and serotonergic receptors and/or transporters in the brain. This work aimed to investigate the cytotoxicity of N‐benzylpiperazine, 1‐(3‐trifluoromethylphenyl)piperazine, 1‐(4‐methoxyphenyl)piperazine and 1‐(3,4‐methylenedioxybenzyl)piperazine in the differentiated human neuroblastoma SH‐SY5Y cell line. Cytotoxicity was evaluated after 24 h incubations through the MTT reduction and neutral red uptake assays. Oxidative stress (reactive oxygen and nitrogen species production and glutathione content) and energetic (ATP content) parameters, as well as intracellular Ca2+, mitochondrial membrane potential, DNA damage (comet assay) and cell death mode were also evaluated. Complete cytotoxicity curves were obtained after 24 h incubations with each drug. A significant decrease in intracellular total glutathione content was noted for all the tested drugs. All drugs caused a significant increase of intracellular free Ca2+ levels, accompanied by mitochondrial hyperpolarization. However, ATP levels remained unchanged. The investigation of cell death mode revealed a predominance of early apoptotic cells. No genotoxicity was found in the comet assay. Among the tested drugs, 1‐(3‐trifluoromethylphenyl)piperazine was the most cytotoxic. Overall, piperazine designer drugs are potentially neurotoxic, supporting concerns on risks associated with the abuse of these drugs. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-20T20:48:22.46823-05:0
      DOI: 10.1002/jat.3153
  • In vitro study of biocompatibility of a graphene composite with gold
           nanoparticles and hydroxyapatite on human osteoblasts
    • Authors: Liana Crisan; Bogdan Crisan, Olga Soritau, Mihaela Baciut, Alexandru Radu Biris, Grigore Baciut, Ondine Lucaciu
      Abstract: The purpose of this study was to evaluate the biocompatibility of some composites consisting of different proportions of graphene in combination with gold nanoparticles (AuNPs) and nanostructured hydroxyapatite (HA) on osteoblast viability, proliferation and differentiation. Au/HA@graphene composites synthesized by the catalytic chemical vapor deposition induction heating method with acetylene as the carbon source and over an Au/HA catalyst, were characterized by transmission electron microscopy, thermogravimetric analysis and Raman spectroscopy and showed that the few‐layer graphene was grown over the Au/HA catalyst. The cytocompatibility study was performed using the fluorescein diacetate assay for assessment of the viability and proliferation of osteoblasts cultivated in the presence of HA, Au/HA and Au/HA@graphene composites as colloidal suspensions or as substrates. The most favorable composites for cell adhesion and proliferation were HA, Au/HA and Au/HA composites with 1.6% and 3.15% concentration of graphenes. Immunocytochemical staining performed after 19 days of osteoblasts cultivation on substrates showed that the graphene composites induced low expression of alkaline phosphatase compared to the control group and HA and Au/HA substrates. The presence of graphene in the substrate composition also induced an increased level of intracellular osteopontin and cytoskeleton reorganization (actin‐F) depending on graphene concentration, suggesting cell activation, increased cellular adhesion and acquisition of a mechanosensorial osteocyte phenotype. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-20T20:15:45.376371-05:
      DOI: 10.1002/jat.3152
  • Preclinical safety evaluation of low molecular weight
           heparin–deoxycholate conjugates as an oral anticoagulant
    • Authors: Ji‐young Kim; Ok‐Cheol Jeon, Hyun Tae Moon, Seung Rim Hwang, Youngro Byun
      Abstract: The preclinical safety of a newly developed oral anticoagulant, the low molecular weight heparin–deoxycholate conjugate (OH09208), was evaluated by a comprehensive evaluating program in compliance with standard guidelines. The single dose oral toxicity study in rats receiving 2000 and 5000 mg kg−1 of OH09208 did not reveal any mortality, unusual body weight changes or necropsy findings. The results of the 4‐week oral toxicity study with a 4‐week recovery program in rats receiving OH09208 in doses of 100, 300 and 1000 mg kg−1 day−1 did not reveal any mortality, or indicate any unusual clinical signs, or show any toxicokinetic relationships to the administration of OH09208. Although the increase in liver enzymes in one male dog treated with 300 mg kg−1 day−1 and one female dog treated with 1000 mg kg−1 day−1 could not be excluded from the effect of the test substance, no other toxicologically significant changes were observed in the 4‐week oral toxicity study with a 4‐week recovery in beagle dogs. Thus, while the no‐observed‐adverse‐effect level value from the 4‐week study in both male and female rats was 1000 mg kg−1 day−1, those from the 4‐week study in male and female beagle dogs were 300 and 1000 mg kg−1 day−1, respectively. Furthermore, OH09208 did not induce anaphylactic reactions in guinea pigs, micronucleated bone marrow cells in male ICR mice, chromosomal aberration in Chinese hamster lung cell lines, bacterial reverse mutation, and any abnormalities in hERG current assay, mouse central nervous system and dog cardiovascular studies. Overall, there were no unexpected toxicities in this preclinical study that might have precluded the safe administration of OH09208 to humans. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-20T19:26:21.800263-05:
      DOI: 10.1002/jat.3146
  • Establishment of a mouse model for amiodarone‐induced liver injury
           and analyses of its hepatotoxic mechanism
    • Authors: Shohei Takai; Shingo Oda, Koichi Tsuneyama, Tatsuki Fukami, Miki Nakajima, Tsuyoshi Yokoi
      Abstract: Drug‐induced liver injury (DILI) is the most frequent cause of post‐marketing warnings and withdrawals. Amiodarone (AMD), an antiarrhythmic, presents a risk of liver injury in humans, and its metabolites, formed by cytochrome P450 3A4, are likely more toxic to hepatocytes than AMD is. However, it remains to be clarified whether the metabolic activation of AMD is involved in liver injury in vivo. In this study, to elucidate the underlying mechanisms of AMD‐induced liver injury, mice were administered AMD [1000 mg kg–1, per os (p.o.)] after pretreatment with dexamethasone [DEX, 60 mg kg–1, intraperitoneal (i.p.)], which induces P450 expression, once daily for 3 days. The plasma alanine aminotransferase (ALT) levels were significantly increased by AMD administration in the DEX‐pretreated mice, and the liver concentrations of desethylamiodarone (DEA), a major metabolite of AMD, were correlated with the changes in the plasma ALT levels. Cytochrome c release into the hepatic cytosol and triglyceride levels in the plasma were increased in DEX plus AMD‐administered mice. Furthermore, the ratio of reduced glutathione to oxidized glutathione disulfide in the liver significantly decreased in the DEX plus AMD‐administered mice. The increase of ALT levels was suppressed by treatment with gadolinium chloride (GdCl3), which is an inhibitor of Kupffer cell function. From these results, it is suggested that AMD and/or DEA contribute to the pathogenesis of AMD‐induced liver injury by producing mitochondrial and oxidative stress and Kupffer cell activation. This study proposes the mechanisms of AMD‐induced liver injury using an in vivo mouse model. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-20T19:19:47.146916-05:
      DOI: 10.1002/jat.3141
  • Bisphenol A inhibits duodenal movement ex vivo of rat through nitric
           oxide‐mediated soluble guanylyl cyclase and α‐adrenergic
           signaling pathways
    • Authors: Kaushik Sarkar; Panchali Tarafder, Goutam Paul
      Abstract: The gastrointestinal tract is directly exposed to bisphenol A (BPA)‐tainted foods and beverages stored in polycarbonate plastic containers. The effect of BPA on the movement of small intestine has not been reported until now. We report here the effect of BPA on the movement of the duodenum ex vivo in a rat model. We found significant inhibition of duodenal movement by BPA (10–320 µ M). We suggest that BPA‐induced inhibition of duodenal movement might be due to the suppression of stimulatory and/or activation of inhibitory motor neurons in enteric plexuses innervating the longitudinal and circular visceral smooth muscle cells in the duodenal wall. We observed a significant reversal of BPA‐induced depression of duodenal movement by methylene blue, a soluble guanylyl cyclase blocker and N‐ω‐nitro‐ L‐arginine methyl ester, a nitric oxide (NO) synthase inhibitor; but significant potentiation of the movement by sodium nitroprusside, a NO donor. From the results, we may suggest that BPA‐induced inhibition of the movement might be partially due to activation of inhibitory motor neurons that secrete NO, a relaxant, on to smooth muscle cells. Furthermore, we found significant reversal of BPA‐induced depression of the movement in phentolamine, an α‐adrenergic receptor blocker, pretreated preparation. This result proves that norepinephrine secreting motor neurons may also be involved in BPA‐induced inhibition of the movement. From the results, we conclude that BPA inhibits the movement of the duodenum through NO‐mediated soluble guanylyl cyclase and α‐adrenergic signaling pathways in visceral smooth muscle cells. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-16T05:40:57.910581-05:
      DOI: 10.1002/jat.3154
  • The surfactant dipalmitoylphophatidylcholine modifies acute responses in
           alveolar carcinoma cells in response to low‐dose silver nanoparticle
    • Authors: A. Murphy; K. Sheehy, A. Casey, G. Chambers
      Abstract: Nanotechnology is a rapidly growing field with silver nanoparticles (AgNP) in particular utilized in a wide variety of consumer products. This has presented a number of concerns relating to exposure and the associated toxicity to humans and the environment. As inhalation is the most common exposure route, this study investigates the potential toxicity of AgNP to A549 alveolar epithelial carcinoma cells and the influence of a major component of lung surfactant dipalmitoylphosphatidylcholine (DPPC) on toxicity. It was illustrated that exposure to AgNP generated low levels of oxidative stress and a reduction in cell viability. While DPPC produced no significant effect on viability studies its presence resulted in increased reactive oxygen species formation. DPPC also significantly modified the inflammatory response generated by AgNP exposure. These findings suggest a possible interaction between AgNP and DPPC causing particles to become more reactive, thus increasing oxidative insult and inflammatory response within A549 cells. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-16T05:34:06.971437-05:
      DOI: 10.1002/jat.3148
  • Microminipigs as a new experimental animal model for toxicological
           studies: comparative pharmacokinetics of perfluoroalkyl acids
    • Authors: Keerthi S. Guruge; Michiko Noguchi, Koji Yoshioka, Eriko Yamazaki, Sachi Taniyasu, Miyako Yoshioka, Noriko Yamanaka, Mitsutaka Ikezawa, Nobuhiko Tanimura, Masumi Sato, Nobuyoshi Yamashita, Hiroaki Kawaguchi
      Abstract: In this study, we evaluated the efficacy of a novel minipig strain, the Microminipig (MMPig), as an animal model for studying the pharmacokinetics of a mixture of 10 perfluoroalkyl acids (PFAAs). After a single oral dose was given, we found that the blood depuration of PFAAs (blood t1/2), which we calculated using first‐order elimination curves, ranged from 1.6 to 86.6 days. Among the five body compartments analyzed, the liver was the greatest site of accumulation of perfluorooctanesulfonate and longer chain perfluorinated carboxylates such as perfluorodecanoic acid, perfluoroundecanoic acid and perfluorododecanoic acid. We observed an increasing accumulation trend of perfluorinated carboxylates in the organs associated with the fluorinated carbon chain length. The perfluorononanoic acid burden was the highest among the treated compounds 21 days after a single exposure, as 29% of the given perfluorononanoic acid dose was accumulated in the tissues. The persistence of PFAAs in edible pig tissues even after 21 days post‐exposure raises concerns about the safety of swine products. This was the first study to use MMPigs to elucidate the pharmacokinetics of a group of environmental pollutants. We found that MMPigs could be excellent experimental animals for toxicological studies due to their easy handling, cost efficacy for target compounds and ease of waste treatment. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-15T08:17:20.50335-05:0
      DOI: 10.1002/jat.3145
  • Acrylamide induces locomotor defects and degeneration of dopamine neurons
           in Caenorhabditis elegans
    • Authors: Jia Li; Dan Li, Yongsheng Yang, Tiantian Xu, Ping Li, Defu He
      Abstract: Acrylamide can form in foods during the cooking process and cause multiple adverse effects. However, the neurotoxicity and mechanisms of acrylamide have not been fully elucidated. In Caenorhabditis elegans, we showed that 48 h exposure to 10–625 mg l−1 acrylamide resulted in a significant decline in locomotor frequency of body bending, head thrashing and pharynx pumping. In addition, acrylamide exposure reduced crawling speeds and changed angles of body bending. It indicates that acrylamide induces locomotor defects, along with parkinsonian‐like movement impairment, including bradykinesia and hypokinesia. Acrylamide also affected chemotaxis plasticity and reduced learning ability. Using transgenic nematodes, we found that acrylamide induced downexpression of Pdat‐1 and led to the degeneration of dopaminergic neurons. Moreover, the enhanced expression of unc‐54, encoding a subunit of α‐synuclein was found. It illustrates that acrylamide is efficient in inducing crucial parkinsonian pathology, including dopaminergic damage and α‐synuclein aggregation. These findings suggest the acrylamide‐induced locomotor defects and neurotoxicity are associated with Parkinson's disease. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-15T07:51:39.378714-05:
      DOI: 10.1002/jat.3144
  • PM2.5‐induced oxidative stress increases adhesion molecules
           expression in human endothelial cells through the
           ERK/AKT/NF‐κB‐dependent pathway
    • Authors: Wei Rui; Longfei Guan, Fang Zhang, Wei Zhang, Wenjun Ding
      Abstract: The aim of this study was to explore the intracellular mechanisms underlying the cardiovascular toxicity of air particulate matter (PM) with an aerodynamic diameter of less than 2.5 µm (PM2.5) in a human umbilical vein cell line, EA.hy926. We found that PM2.5 exposure triggered reactive oxygen species (ROS) generation, resulting in a significant decrease in cell viability. Data from Western blots showed that PM2.5 induced phosphorylation of Jun N‐terminal kinase (JNK), extracellular signal regulatory kinase (ERK), p38 mitogen‐activated protein kinase (MAPK) and protein kinase B (AKT), and activation of nuclear factor kappa B (NF‐κB). We further observed a significant increase in expressions of intercellular adhesion molecule‐1 (ICAM‐1) and vascular adhesion molecule‐1 (VCAM‐1) in a time‐ and dose‐dependent manner. Moreover, the adhesion of monocytic THP‐1 cells to EA.hy926 cells was greatly enhanced in the presence of PM2.5. However, N‐acetylcysteine (NAC), a scavenger of ROS, prevented the increase of ROS generation, attenuated the phosphorylation of the above kinases, and decreased the NF‐κB activation as well as the expression of ICAM‐1 and VCAM‐1. Furthermore, ERK inhibitor (U0126), AKT inhibitor (LY294002) and NF‐κB inhibitor (BAY11‐7082) significantly down‐regulated PM2.5‐induced ICAM‐1 and VCAM‐1 expression as well as adhesion of THP‐1 cells, but not JNK inhibitor (SP600125) and p38 MAPK inhibitor (SB203580), indicating that ERK/AKT/NF‐κB is involved in the signaling pathway that leads to PM2.5‐induced ICAM‐1 and VCAM‐1 expression. These findings suggest PM2.5‐induced ROS may function as signaling molecules triggering ICAM‐1 and VCAM‐1 expressions through activating the ERK/AKT/NF‐κB‐dependent pathway, and further promoting monocyte adhesion to endothelial cells. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-15T07:37:09.723701-05:
      DOI: 10.1002/jat.3143
  • Aluminium oxide nanoparticles induced morphological changes, cytotoxicity
           and oxidative stress in Chinook salmon (CHSE‐214) cells
    • Authors: Koigoora Srikanth; Amit Mahajan, Eduarda Pereira, Armando Costa Duarte, Janapala Venkateswara Rao
      Abstract: Aluminium oxide nanoparticles (Al2O3 NPs) are increasingly used in diverse applications that has raised concern about their safety. Recent studies suggested that Al2O3 NPs induced oxidative stress may be the cause of toxicity in algae, Ceriodaphnia dubia, Caenorhabditis elegans and Danio rerio. However, there is paucity on the toxicity of Al2O3 NPs on fish cell lines. The current study was aimed to investigate Al2O3 NPs induced cytotoxicity, oxidative stress and morphological abnormality of Chinnok salmon cells (CHSE‐214). A dose‐dependent decline in cell viability was observed in CHSE‐214 cells exposed to Al2O3 NPs. Oxidative stress induced by Al2O3 NPs in CHSE‐214 cells has resulted in the significant reduction of superoxide dismutase, catalase and glutathione in a dose‐dependent manner. However, a significant increase in glutathione sulfo‐transferase and lipid peroxidation was observed in CHSE‐214 cells exposed to Al2O3 NPs in a dose‐dependent manner. Significant morphological changes in CHSE‐214 cells were observed when exposed to Al2O3 NPs at 6, 12 and 24 h. The cells started to detach and appear spherical at 6 h followed by loss of cellular contents resulting in the shrinking of the cells. At 24 h, the cells started to disintegrate and resulted in cell death. Our data demonstrate that Al2O3 NPs induce cytotoxicity and oxidative stress in a dose‐dependent manner in CHSE‐214 cells. Thus, our current work may serve as a base‐line study for future evaluation of toxicity studies using CHSE‐214 cells. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-15T07:06:31.862304-05:
      DOI: 10.1002/jat.3142
  • Particulate matter phagocytosis induces tissue factor in differentiating
    • Authors: M. Milano; P. Dongiovanni, A. Artoni, S. Gatti, L. Rosso, F. Colombo, V. Bollati, M. Maggioni, P. M. Mannucci, P. A. Bertazzi, S. Fargion, L. Valenti
      Abstract: Airborne exposure to particulate matter with diameter
      PubDate: 2015-04-08T22:41:25.814999-05:
      DOI: 10.1002/jat.3156
  • Impaired aquaporins expression in the gastrointestinal tract of rat after
           mercury exposure
    • Authors: Cinzia Bottino; Marta Vázquez, Vicenta Devesa, Umberto Laforenza
      Abstract: The main route of exposure to mercury in humans is through the diet. Consequently, the gastrointestinal mucosa is exposed to the mercurial forms, where they cause intestinal fluid accumulation, mucosal injuries and diarrhea. The relationship between inorganic mercury (HgCl2) and methylmercury (CH3HgCl) exposure and water movement in the gastrointestinal tract is still unexplored. The leading role of aquaporins (AQPs) in the rapid bidirectional movement of fluid in the gastrointestinal tract of mammals is well established. The present study evaluates the effect of HgCl2 and CH3HgCl exposure on AQP expression in different portions of the gastrointestinal tract of rats treated by gavage (5 mg kg–1 of mercury species, single dose, 4 days). The results show that mercury species reduce mRNA and protein levels of AQPs in different parts of the gastrointestinal tract. In the stomach, treated rats show a significant reduction of expression of AQP3 (80–90% for mRNA and 50% for protein) and AQP4 (95–99% for mRNA and 20–40% for protein). In the small and large intestine, treated rats experience a significant reduction of AQP3 and AQP7 expression. Protein contents of both AQPs are reduced in similar proportions in jejunum (AQP3: 40–50%; AQP7: 45–50%) and colon (AQP3: 35–40%; AQP7: 45–60%), regardless of the treatment. Our results indicate that some AQPs are downregulated in the rat gastrointestinal tract by mercury exposure, suggesting a possible role of AQPs in the development of mercury gastrointestinal symptoms. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-08T20:55:50.964966-05:
      DOI: 10.1002/jat.3151
  • Co‐treatment with the non‐steroidal anti‐androgen drug,
           flutamide and the natural estrogen, 17β‐estradiol does not lead
           to additive reproductive impairment in juvenile Murray rainbowfish
           (Melanotaenia fluviatilis)
    • Authors: Harpreet Bhatia; Anupama Kumar, Jun Du, John C. Chapman, Mike J. McLaughlin
      Abstract: The aim of this study was to investigate if the anti‐androgen, flutamide, and the estrogen, 17β‐estradiol work together to feminize juvenile Murray rainbowfish (Melanotaenia fluviatilis). Fish (60 days post‐hatch) were exposed to 25 ng/L 17β‐estradiol (E2), 25 µg/L flutamide (Flu low), 250 µg/L flutamide (Flu high), E2 + Flu low and E2 + Flu high. After 35 days of exposure, concentrations of sex steroid hormones, 17β‐estradiol and 11‐keto testosterone (11‐KT), were determined in the head; and vitellogenin (VTG) concentration was measured in the tail. The abdomens were used for histological investigation of the gonads. Treatment with E2 + Flu high resulted in reduction in body weights and lengths in males and condition factor in females. Intersex was noted in Flu high and E2 + Flu high treatments. Exposures to E2 and/or Flu (low and high) resulted in precocious oocyte development but inhibited sperm development. The 17β‐estradiol levels decreased significantly in the heads of both sexes after exposures to E2 and/or Flu (high and low). Flu high and E2 alone increased the 11‐KT levels in both sexes. However, E2 + Flu low decreased 11‐KT levels in males and increased them in females. Flutamide (low and high) induced VTG protein in the tails of both sexes. In males, VTG was not induced in the tail after exposure to E2. No significant effect of flutamide on E2‐induced VTG concentration was noted. We conclude that co‐treatment with flutamide and 17β‐estradiol does not lead to additive reproductive impairment in juvenile Murray rainbowfish. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-04-08T07:40:51.66153-05:0
      DOI: 10.1002/jat.3135
  • Titanium dioxide nanoparticles increase plasma glucose via reactive oxygen
           species‐induced insulin resistance in mice
    • Authors: Hailong Hu; Qian Guo, Changlin Wang, Xiao Ma, Hongjuan He, Yuri Oh, Yujie Feng, Qiong Wu, Ning Gu
      Abstract: There have been few reports about the possible toxic effects of titanium dioxide (TiO2) nanoparticles on the endocrine system. We explored the endocrine effects of oral administration to mice of anatase TiO2 nanoparticles (0, 64 and 320 mg kg–1 body weight per day to control, low‐dose and high‐dose groups, respectively, 7 days per week for 14 weeks). TiO2 nanoparticles were characterized by scanning and transmission electron microscopy (TEM) and dynamic light scattering (DLS), and their physiological distribution was investigated by inductively coupled plasma. Biochemical analyzes included plasma glucose, insulin, heart blood triglycerides (TG), free fatty acid (FFA), low‐density lipoprotein cholesterol (LDL‐C), high‐density lipoprotein cholesterol (HDL‐C), total cholesterol (TC), tumor necrosis factor‐alpha (TNF‐α), interleukin (IL)‐6 and reactive oxygen species (ROS)‐related markers (total SOD, GSH and MDA). Phosphorylation of IRS1, Akt, JNK1, and p38 MAPK were analyzed by western blotting. Increased titanium levels were found in the liver, spleen, small intestine, kidney and pancreas. Biochemical analyzes showed that plasma glucose significantly increased whereas there was no difference in plasma insulin secretion. Increased ROS levels were found in serum and the liver, as evidenced by reduced total SOD activity and GSH level and increased MDA content. Western blotting showed that oral administration of TiO2 nanoparticles induced insulin resistance (IR) in mouse liver, shown by increased phosphorylation of IRS1 (Ser307) and reduced phosphorylation of Akt (Ser473). The pathway by which TiO2 nanoparticles increase ROS‐induced IR were included in the inflammatory response and phosphokinase, as shown by increased serum levels of TNF‐α and IL‐6 and increased phosphorylation of JNK1 and p38 MAPK in liver. These results show that oral administration of TiO2 nanoparticles increases ROS, resulting in IR and increasing plasma glucose in mice. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-31T05:59:02.869333-05:
      DOI: 10.1002/jat.3150
  • Augmenting effects of gestational arsenite exposure of C3H mice on the
           hepatic tumors of the F2 male offspring via the F1 male offspring
    • Authors: Keiko Nohara; Kazuyuki Okamura, Takehiro Suzuki, Hikari Murai, Takaaki Ito, Keiko Shinjo, Shota Takumi, Takehiro Michikawa, Yutaka Kondo, Kenichiro Hata
      Abstract: Gestational exposure can affect the F2 generation through exposure of F1 germline cells. Previous studies reported that arsenite exposure of only F0 females during their pregnancy increases hepatic tumors in the F1 males in C3H mice, whose males are predisposed spontaneously to develop hepatic tumors later in life. The present study addressed the effects of gestational arsenite exposure on tumorigenesis of the F2 males in C3H mice. Expression analysis of several genes in the normal livers at 53 and 80 weeks of age clearly showed significant changes in the F2 males obtained by crossing gestational arsenite‐exposed F1 (arsenite‐F1) males and females compared to the control F2 males. Some of the changes were shown to occur in a late‐onset manner. Then the tumor incidence was assessed at 75–82 weeks of age in the F2 males obtained by reciprocal crossing between the control and arsenite‐F1 males and females. The results demonstrated that the F2 males born to arsenite‐F1 males developed tumors at a significantly higher rate than the F2 males born to the control F1 males, irrespective of exposure of F1 females. Gene expressions of hepatocellular carcinoma markers β‐catenin (CTNNB1) and interleukin‐1 receptor antagonist in the tumors were significantly upregulated in the F2 males born to arsenite‐F1 males compared to those born to the control F1 males. These results show that arsenite exposure of only F0 pregnant mice causes late‐onset changes and augments tumors in the livers of the F2 males by affecting the F1 male offspring. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-30T22:47:14.094101-05:
      DOI: 10.1002/jat.3149
  • Gene expression profiling of the hippocampal dentate gyrus in an adult
           toxicity study captures a variety of neurodevelopmental dysfunctions in
           rat models of hypothyroidism
    • Authors: Ayako Shiraki; Fumiyo Saito, Hirotoshi Akane, Yumi Akahori, Nobuya Imatanaka, Megu Itahashi, Toshinori Yoshida, Makoto Shibutani
      Abstract: We previously found that developmental hypothyroidism changed the expression of genes in the rat hippocampal dentate gyrus, a brain region where adult neurogenesis is known to occur. In the present study, we performed brain region‐specific global gene expression profiling in an adult rat hypothyroidism model to see if it reflected the developmental neurotoxicity we saw in the developmental hypothyroidism model. Starting when male rats were 5 weeks old, we administered 6‐propyl‐2‐thiouracil at a doses of 0, 0.1 and 10 mg kg−1 body weight by gavage for 28 days. We selected four brain regions to represent both cerebral and cerebellar tissues: hippocampal dentate gyrus, cerebral cortex, corpus callosum and cerebellar vermis. We observed significant alterations in the expression of genes related to neural development (Eph family genes and Robo3) in the cerebral cortex and hippocampal dentate gyrus and in the expression of genes related to myelination (Plp1 and Mbp) in the hippocampal dentate gyrus. We observed only minor changes in the expression of these genes in the corpus callosum and cerebellar vermis. We used real‐time reverse‐transcription polymerase chain reaction to confirm Chrdl1, Hes5, Mbp, Plp1, Slit1, Robo3 and the Eph family transcript expression changes. The most significant changes in gene expression were found in the dentate gyrus. Considering that the gene expression profile of the adult dentate gyrus closely related to neurogenesis, 28‐day toxicity studies looking at gene expression changes in adult hippocampal dentate gyrus may also detect possible developmental neurotoxic effects. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-30T20:50:06.345358-05:
      DOI: 10.1002/jat.3140
  • Development of toxicity values and exposure estimates for
           tetrabromobisphenol A: application in a margin of exposure assessment
    • Authors: Daniele Wikoff; Chad Thompson, Camarie Perry, Matthew White, Susan Borghoff, Lauren Fitzgerald, Laurie C. Haws
      Abstract: Tetrabromobisphenol A (TBBPA) is used in a diverse array of products to improve fire safety. The National Toxicology Program (NTP) recently completed a 2‐year bioassay for TBBPA. The objective of the present study was to develop a cancer‐based and a non‐cancer based toxicity value and to compare such to appropriate estimates of human exposure. Data from the NTP 2‐year and 13‐week studies were selected to develop candidate toxicity values. Benchmark dose modeling and subsequent evaluation of candidate values resulted in selection of an oral reference dose (RfD) of 0.6 mg kg−1 day−1 based on uterine hyperplasia in rats and an oral cancer slope factor (OSF) of 0.00315 per mg kg−1 day−1 based on an increased incidence of uterine tumors in rats. Lifetime average daily dose (LADD) estimates ranged from 2.2 E−7 to 3.9 E−6 mg kg−1 day−1 based on age‐adjusted exposures to TBBPA via breast milk consumption, dietary intake, soil/dust ingestion and drinking water ingestion in infants, young children, older children and adults. Average daily dose (ADD) estimates ranged from 3.2 E −7 to 8.4 E−5 mg kg−1 day−1. Resulting margin of exposure (MOE) values were > 800 000 for non‐cancer endpoints and > 32 000 000 for cancer‐based endpoints. These data collectively indicate a low level of health concern associated with exposures to TBBPA based on current data. It is anticipated that the exposure estimates, along with the toxicity values described within, should be informative for understanding human health hazards associated with TBBPA. © 2015. The
      Authors . Journal of Applied Toxicology Published by John Wiley & Sons Ltd.
      PubDate: 2015-03-30T20:10:22.288627-05:
      DOI: 10.1002/jat.3132
  • Evaluation of combinations of in vitro sensitization test descriptors for
           the artificial neural network‐based risk assessment model of skin
    • Authors: Morihiko Hirota; Shiho Fukui, Kenji Okamoto, Satoru Kurotani, Noriyasu Imai, Miyuki Fujishiro, Daiki Kyotani, Yoshinao Kato, Toshihiko Kasahara, Masaharu Fujita, Akemi Toyoda, Daisuke Sekiya, Shinichi Watanabe, Hirokazu Seto, Osamu Takenouchi, Takao Ashikaga, Masaaki Miyazawa
      Abstract: The skin sensitization potential of chemicals has been determined with the use of the murine local lymph node assay (LLNA). However, in recent years public concern about animal welfare has led to a requirement for non‐animal risk assessment systems for the prediction of skin sensitization potential, to replace LLNA. Selection of an appropriate in vitro test or in silico model descriptors is critical to obtain good predictive performance. Here, we investigated the utility of artificial neural network (ANN) prediction models using various combinations of descriptors from several in vitro sensitization tests. The dataset, collected from published data and from experiments carried out in collaboration with the Japan Cosmetic Industry Association (JCIA), consisted of values from the human cell line activation test (h‐CLAT), direct peptide reactivity assay (DPRA), SH test and antioxidant response element (ARE) assay for chemicals whose LLNA thresholds have been reported. After confirming the relationship between individual in vitro test descriptors and the LLNA threshold (e.g. EC3 value), we used the subsets of chemicals for which the requisite test values were available to evaluate the predictive performance of ANN models using combinations of h‐CLAT/DPRA (N = 139 chemicals), the DPRA/ARE assay (N = 69), the SH test/ARE assay (N = 73), the h‐CLAT/DPRA/ARE assay (N = 69) and the h‐CLAT/SH test/ARE assay (N = 73). The h‐CLAT/DPRA, h‐CLAT/DPRA/ARE assay and h‐CLAT/SH test/ARE assay combinations showed a better predictive performance than the DPRA/ARE assay and the SH test/ARE assay. Our data indicates that the descriptors evaluated in this study were all useful for predicting human skin sensitization potential, although combinations containing h‐CLAT (reflecting dendritic cell‐activating ability) were most effective for ANN‐based prediction. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-30T19:40:57.244347-05:
      DOI: 10.1002/jat.3105
  • Test battery with the human cell line activation test, direct peptide
           reactivity assay and DEREK based on a 139 chemical data set for predicting
           skin sensitizing potential and potency of chemicals
    • Authors: Osamu Takenouchi; Shiho Fukui, Kenji Okamoto, Satoru Kurotani, Noriyasu Imai, Miyuki Fujishiro, Daiki Kyotani, Yoshinao Kato, Toshihiko Kasahara, Masaharu Fujita, Akemi Toyoda, Daisuke Sekiya, Shinichi Watanabe, Hirokazu Seto, Morihiko Hirota, Takao Ashikaga, Masaaki Miyazawa
      Abstract: To develop a testing strategy incorporating the human cell line activation test (h‐CLAT), direct peptide reactivity assay (DPRA) and DEREK, we created an expanded data set of 139 chemicals (102 sensitizers and 37 non‐sensitizers) by combining the existing data set of 101 chemicals through the collaborative projects of Japan Cosmetic Industry Association. Of the additional 38 chemicals, 15 chemicals with relatively low water solubility (log Kow > 3.5) were selected to clarify the limitation of testing strategies regarding the lipophilic chemicals. Predictivities of the h‐CLAT, DPRA and DEREK, and the combinations thereof were evaluated by comparison to results of the local lymph node assay. When evaluating 139 chemicals using combinations of three methods based on integrated testing strategy (ITS) concept (ITS‐based test battery) and a sequential testing strategy (STS) weighing the predictive performance of the h‐CLAT and DPRA, overall similar predictivities were found as before on the 101 chemical data set. An analysis of false negative chemicals suggested a major limitation of our strategies was the testing of low water‐soluble chemicals. When excluded the negative results for chemicals with log Kow > 3.5, the sensitivity and accuracy of ITS improved to 97% (91 of 94 chemicals) and 89% (114 of 128). Likewise, the sensitivity and accuracy of STS to 98% (92 of 94) and 85% (111 of 129). Moreover, the ITS and STS also showed good correlation with local lymph node assay on three potency classifications, yielding accuracies of 74% (ITS) and 73% (STS). Thus, the inclusion of log Kow in analysis could give both strategies a higher predictive performance. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-29T21:26:10.794897-05:
      DOI: 10.1002/jat.3127
  • A novel in chemico method to detect skin sensitizers in highly diluted
           reaction conditions
    • Authors: Yusuke Yamamoto; Haruna Tahara, Ryota Usami, Toshihiko Kasahara, Yoshihiro Jimbo, Takanori Hioki, Masaharu Fujita
      Abstract: The direct peptide reactivity assay (DPRA) is a simple and versatile alternative method for the evaluation of skin sensitization that involves the reaction of test chemicals with two peptides. However, this method requires concentrated solutions of test chemicals, and hydrophobic substances may not dissolve at the concentrations required. Furthermore, hydrophobic test chemicals may precipitate when added to the reaction solution. We previously established a high‐sensitivity method, the amino acid derivative reactivity assay (ADRA). This method uses novel cysteine (NAC) and novel lysine derivatives (NAL), which were synthesized by introducing a naphthalene ring to the amine group of cysteine and lysine residues. In this study, we modified the ADRA method by reducing the concentration of the test chemicals 100‐fold. We investigated the accuracy of skin sensitization predictions made using the modified method, which was designated the ADRA‐dilutional method (ADRA‐DM). The predictive accuracy of the ADRA‐DM for skin sensitization was 90% for 82 test chemicals which were also evaluated via the ADRA, and the predictive accuracy in the ADRA‐DM was higher than that in the ADRA and DPRA. Furthermore, no precipitation of test compounds was observed at the initiation of the ADRA‐DM reaction. These results show that the ADRA‐DM allowed the use of test chemicals at concentrations two orders of magnitude lower than that possible with the ADRA. In addition, ADRA‐DM does not have the restrictions on test compound solubility that were a major problem with the DPRA. Therefore, the ADRA‐DM is a versatile and useful method. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-25T00:19:24.155741-05:
      DOI: 10.1002/jat.3139
  • Particle uptake efficiency is significantly affected by type of capping
           agent and cell line
    • Authors: Fan Zhang; Phillip Durham, Christie M. Sayes, Boris L. T. Lau, Erica D. Bruce
      Abstract: Surface‐functionalized silver nanoparticles (AgNPs) are the most deployed engineered nanomaterials in consumer products because of their optical, antibacterial and electrical properties. Almost all engineered nanoparticles are coated with application‐specific capping agents (i.e. organic/inorganic ligands on particle surface) to enhance their stability in suspension or increase their biocompatibility for biomedicine. The aim of this study was to investigate the contribution of the selected capping agents to their observed health impacts using realistic dose ranges. AgNPs capped with citrate, polyvinylpyrrolidone (PVP) and tannic acid were studied with human bronchoalveolar carcinoma (A549) and human colon adenocarcinoma (Caco‐2) cell lines and compared against exposures to Ag ions. Cellular uptake and cytotoxicity were evaluated up to 24 h. Tannic acid capped AgNPs induced higher cellular uptake and rate in both cell lines. Citrate‐capped and PVP‐capped AgNPs behaved similarly over 24 h. All three of the capped AgNPs penetrated more into the A549 cells than Caco‐2 cells. In contrast, the uptake rate of Ag ions in Caco‐2 cells (0.11 ± 0.0001 µg h–1) was higher than A549 cells (0.025 ± 0.00004 µg h–1). The exposure concentration of 3 mg l–1 is below the EC50 value for all of the AgNPs; therefore, little cytotoxicity was observed in any experiment conducted herein. Exposure of Ag ions, however, interrupted cell membrane integrity and cell proliferation (up to 70% lysed after 24 h). These findings indicate cellular uptake is dependent on capping agent, and when controlled to realistic exposure concentrations, cellular function is not significantly affected by AgNP exposure. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-24T23:21:44.462642-05:
      DOI: 10.1002/jat.3138
  • Comparative effects of sulfhydryl compounds on target organellae, nuclei
           and mitochondria, of hydroxylated fullerene‐induced cytotoxicity in
           isolated rat hepatocytes
    • Authors: Yoshio Nakagawa; Akiko Inomata, Akio Ogata, Dai Nakae
      Abstract: DNA damage and cytotoxicity induced by a hydroxylated fullerene [C60(OH)24], which is a spherical nanomaterial and/or a water‐soluble fullerene derivative, and their protection by sulfhydryl compounds were studied in freshly isolated rat hepatocytes. The exposure of hepatocytes to C60(OH)24 at a concentration of 50 μM caused time (0 to 3 h)‐dependent cell death accompanied by the formation of cell surface blebs, the loss of cellular levels of ATP and reduced glutathione, accumulation of glutathione disulfide, and induction of DNA fragmentation assayed using alkali single‐cell agarose‐gel electrophoresis. C60(OH)24‐induced cytotoxicity was effectively prevented by pretreatment with sulfhydryl compounds. N‐acetyl‐L‐cysteine (NAC), L‐cysteine and L‐methionine, at a concentration of 2.5 mM, ameliorated cell death, accompanied by a decrease in cellular ATP levels, formation of cell surface blebs, induction of reactive oxygen species (ROS) and loss of mitochondrial membrane potential caused by C60(OH)24. In addition, DNA fragmentation caused by C60(OH)24 was also inhibited by NAC, whereas an antioxidant ascorbic acid did not affect C60(OH)24‐induced cell death and DNA damage in rat hepatocytes. Taken collectively, these results indicate that incubation of rat hepatocytes with C60(OH)24 elicits DNA damage, suggesting that nuclei as well as mitochondria are target sites of the hydroxylated fullerene; and induction of DNA damage and oxidative stress is ameliorated by an increase in cellular GSH levels, suggesting that the onset of toxic effects may be partially attributable to a thiol redox‐state imbalance caused by C60(OH)24. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-23T04:41:18.036497-05:
      DOI: 10.1002/jat.3137
  • Meloxicam inhibits fipronil‐induced apoptosis via modulation of the
           oxidative stress and inflammatory response in SH‐SY5Y cells
    • Authors: Jae Hyeon Park; Youn Sun Park, Je‐Bong Lee, Kyung‐Hun Park, Min‐kyoung Paik, Mihye Jeong, Hyun Chul Koh
      Abstract: Oxidative stress and inflammatory responses have been identified as key elements of neuronal cell apoptosis. In this study, we investigated the mechanisms by which inflammatory responses contribute to apoptosis in human neuroblastoma SH‐SY5Y cells treated with fipronil (FPN). Based on the cytotoxic mechanism of FPN, we examined the neuroprotective effects of meloxicam against FPN‐induced neuronal cell death. Treatment of SH‐SY5Y cells with FPN induced apoptosis via activation of caspase‐9 and ‐3, leading to nuclear condensation. In addition, FPN induced oxidative stress and increased expression of cyclooxygenase‐2 (COX‐2) and tumor necrosis factor‐α (TNF‐α) via inflammatory stimulation. Pretreatment of cells with meloxicam enhanced the viability of FPN‐exposed cells through attenuation of oxidative stress and inflammatory response. FPN activated mitogen activated protein kinase (MAPK) and inhibitors of MAPK abolished FPN‐induced COX‐2 expression. Meloxicam also attenuated FPN‐induced cell death by reducing MAPK‐mediated pro‐inflammatory factors. Furthermore, we observed both nuclear accumulation of p53 and enhanced levels of cytosolic p53 in a concentration‐dependent manner after FPN treatment. Pretreatment of cells with meloxicam blocked the translocation of p53 from the cytosol to the nucleus. Together, these data suggest that meloxicam may exert anti‐apoptotic effects against FPN‐induced cytotoxicity by both attenuating oxidative stress and inhibiting the inflammatory cascade via inactivation of MAPK and p53 signaling. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-13T12:18:27.333032-05:
      DOI: 10.1002/jat.3136
  • Investigation on cobalt‐oxide nanoparticles cyto‐genotoxicity
           and inflammatory response in two types of respiratory cells
    • Authors: Delia Cavallo; Aureliano Ciervo, Anna Maria Fresegna, Raffaele Maiello, Paola Tassone, Giuliana Buresti, Stefano Casciardi, Sergio Iavicoli, Cinzia Lucia Ursini
      Abstract: The increasing use of cobalt oxide (Co3O4) nanoparticles (NPs) in several applications and the suggested genotoxic potential of Co‐oxide highlight the importance of evaluating Co3O4 NPs toxicity. Cyto‐genotoxic and inflammatory effects induced by Co3O4 NPs were investigated in human alveolar (A549), and bronchial (BEAS‐2B) cells exposed to 1–40 µg ml–1. The physicochemical properties of tested NPs were analysed by transmission electron microscopy (TEM) and dynamic light scattering (DLS). Cytotoxicity was studied to analyze cell viability (WST1 test) and membrane damage (LDH assay), direct/oxidative DNA damage was assessed by the Formamido‐pyrimidine glycosylase (Fpg)‐modified comet assay and inflammation by interleukin (IL)‐6, IL‐8 and tumor necrosis factor‐alpha (TNF‐α) release (ELISA). In A549 cells, no cytotoxicity was found, whereas BEAS‐2B cells showed a viability reduction at 40 µg ml–1 and early membrane damage at 1, 5 and 40 µg ml–1. In A549 cells, direct and oxidative DNA damage at 20 and 40 µg ml–1 were detected without any effects on cytokine release. In BEAS‐2B cells, significant direct DNA damage at 40 µg ml–1 and significant oxidative DNA damage with a peak at 5 µg ml–1, that was associated with increased TNF‐α release at 1 µg ml–1 after 2 h and increased IL‐8 release at 20 µg ml–1 after 24 h, were detected. The findings show in the transformed alveolar cells no cytotoxicity and genotoxic/oxidative effects at 20 and 40 µg ml–1. In normal bronchial cells, moderate cytotoxicity, direct DNA damage only at the highest concentration and significant oxidative‐inflammatory effects at lower concentrations were detected. The findings confirm the genotoxic‐oxidative potential of Co3O4 NPs and show greater sensitivity of BEAS‐2B cells to cytotoxic and oxidative‐inflammatory effects suggesting the use of different cell lines and multiple end‐points to elucidate Co3O4 NPs toxicity. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-13T12:08:33.285112-05:
      DOI: 10.1002/jat.3133
  • Cellular localization of uranium in the renal proximal tubules during
           acute renal uranium toxicity
    • Authors: Shino Homma‐Takeda; Keisuke Kitahara, Kyoko Suzuki, Benjamin J. Blyth, Noriyoshi Suya, Teruaki Konishi, Yasuko Terada, Yoshiya Shimada
      Abstract: Renal toxicity is a hallmark of uranium exposure, with uranium accumulating specifically in the S3 segment of the proximal tubules causing tubular damage. As the distribution, concentration and dynamics of accumulated uranium at the cellular level is not well understood, here, we report on high‐resolution quantitative in situ measurements by high‐energy synchrotron radiation X‐ray fluorescence analysis in renal sections from a rat model of uranium‐induced acute renal toxicity. One day after subcutaneous administration of uranium acetate to male Wistar rats at a dose of 0.5 mg uranium kg–1 body weight, uranium concentration in the S3 segment of the proximal tubules was 64.9 ± 18.2 µg g–1, sevenfold higher than the mean renal uranium concentration (9.7 ± 2.4 µg g–1). Uranium distributed into the epithelium of the S3 segment of the proximal tubules and highly concentrated uranium (50‐fold above mean renal concentration) in micro‐regions was found near the nuclei. These uranium levels were maintained up to 8 days post‐administration, despite more rapid reductions in mean renal concentration. Two weeks after uranium administration, damaged areas were filled with regenerating tubules and morphological signs of tissue recovery, but areas of high uranium concentration (100‐fold above mean renal concentration) were still found in the epithelium of regenerating tubules. These data indicate that site‐specific accumulation of uranium in micro‐regions of the S3 segment of the proximal tubules and retention of uranium in concentrated areas during recovery are characteristics of uranium behavior in the kidney. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-13T11:52:31.309239-05:
      DOI: 10.1002/jat.3126
  • Time profiles and toxicokinetic parameters of key biomarkers of exposure
           to cypermethrin in orally exposed volunteers compared with previously
           available kinetic data following permethrin exposure
    • Authors: Mylène Ratelle; Jonathan Coté, Michèle Bouchard
      Abstract: Biomonitoring of pyrethroid exposure is largely conducted but human toxicokinetics has not been fully documented. This is essential for a proper interpretation of biomonitoring data. Time profiles and toxicokinetic parameters of key biomarkers of exposure to cypermethrin in orally exposed volunteers have been documented and compared with previously available kinetic data following permethrin dosing. Six volunteers ingested 0.1 mg kg–1 bodyweight of cypermethrin acutely. The same volunteers were exposed to permethrin earlier. Blood samples were taken over 72 h after treatment and complete timed urine voids were collected over 84 h postdosing. Cis‐ and trans‐3‐(2,2‐dichlorovinyl)‐2,2‐dimethylcyclopropane‐1‐carboxylic acids (trans‐ and cis‐DCCA) and 3‐phenoxybenzoic acid (3‐PBA) metabolites, common to both cypermethrin and permethrin, were quantified. Blood and urinary time courses of all three metabolites were similar following cypermethrin and permethrin exposure. Plasma levels of metabolites reached peak values on average ≈ 5–7 h post‐dosing; the elimination phase showed mean apparent half‐lives (t½) for trans‐DCCA, cis‐DCCA and 3‐PBA of 5.1, 6.9 and 9.2 h, respectively, following cypermethrin treatment as compared to 7.1, 6.2 and 6.5 h after permethrin dosing. Corresponding mean values obtained from urinary rate time courses were peak values at ≈ 9 h post‐dosing and apparent elimination t½ of 6.3, 6.4 and 6.4 h for trans‐DCCA, cis‐DCCA and 3‐PBA, respectively, following cypermethrin treatment as compared to 5.4, 4.5 and 5.7 h after permethrin dosing. These data confirm that the kinetics of cypermethrin is similar to that of permethrin in humans and that their common biomarkers of exposure may be used for an overall assessment of exposure. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-13T11:33:30.932661-05:
      DOI: 10.1002/jat.3124
  • Non‐clinical safety assessment of single and repeated intramuscular
           administration of a human papillomavirus‐16/18 vaccine in rabbits
           and rats
    • Authors: Lawrence Segal; Danielle Morelle, Kari Kaaber, Eric Destexhe, Nathalie Garçon
      Abstract: The human papillomavirus (HPV)‐16/18 vaccine (Cervarix®) is a prophylactic vaccine for the prevention of cervical cancer. The vaccine contains recombinant virus‐like particles assembled from the L1 major capsid proteins of the cervical cancer‐causing viral types HPV‐16 and HPV‐18, and Adjuvant System 04 (AS04), which contains the immunostimulant MPL and aluminium salt. To evaluate potential local and systemic toxic effects of the HPV‐16/18 vaccine or AS04 alone, three repeated‐dose studies were performed in rabbits and rats. One rabbit study also included a single‐dose evaluation. In rabbits (~2.5 kg), the full human dose (HD) of the vaccine was evaluated (0.5 ml per injection site), and in rats (~250 g), 1/5 HD of vaccine was evaluated, corresponding to ≥ 12 times the dosage in humans relative to body weight. In both animal models, the treatment‐related changes included a slight transient increase in the number of circulating neutrophils as well as a local inflammatory reaction at the injection site. These treatment‐related changes were less pronounced after four doses of AS04 alone than after four doses of the HPV‐16/18 vaccine. Additional treatment‐related changes in the rat included lower albumin/globulin ratios and microscopic signs of inflammation in the popliteal lymph nodes. In both animal models, 13 weeks after the fourth dose, recovery was nearly complete, although at the injection site in some animals there were signs of discoloration, muscle‐fibre regeneration and focal points of macrophage infiltration. Therefore, in these non‐clinical models, the single and repeated dose administrations of the HPV‐16/18 vaccine or AS04 alone were safe and well tolerated. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-06T09:02:51.11051-05:0
      DOI: 10.1002/jat.3131
  • Non‐clinical safety and biodistribution of AS03‐adjuvanted
           inactivated pandemic influenza vaccines
    • Authors: Lawrence Segal; Sandrine Wouters, Danielle Morelle, Gaëlle Gautier, Julien Le Gal, Thomas Martin, Frieke Kuper, Eric Destexhe, Arnaud M. Didierlaurent, Nathalie Garçon
      Abstract: Pandemic‐influenza vaccines containing split‐inactivated‐virus antigen have been formulated with the immunostimulatory Adjuvant System AS03 to enhance the antigen immunogenicity and reduce antigen content per dose. AS03 is an oil‐in‐water emulsion containing α‐tocopherol, squalene and polysorbate 80. To support the clinical development of AS03‐adjuvanted pandemic‐influenza vaccines, the local and systemic toxicity of test articles containing split‐influenza A(H5N1) and/or AS03 were evaluated after 3–4 intramuscular (i.m.) injections in rabbits. Treatment‐related effects were restricted to mild inflammatory responses and were induced primarily by the test articles containing AS03. The injection‐site inflammation was mild at 3 days, and minimal at 4 weeks after the last injection; and was reflected by signs of activation in the draining lymph nodes and by systemic effects in the blood including a transient increase of neutrophils. In addition, a study in mice explored the biodistribution of A(H5N1) vaccines or AS03 through radiolabelling the antigen or constituents of AS03 prior to injection. In this evaluation, 57–73% of AS03's principal constituents had cleared from the injection site 3 days after injection, and their different clearance kinetics were suggestive of AS03's dissociation. All these AS03 constituents entered into the draining lymph nodes within 30 min after injection. In conclusion, the administration of repeated doses of the H5N1/AS03 vaccine was well tolerated in the rabbit, and was primarily associated with transient mild inflammation at the injection site and draining lymph nodes. The biodistribution kinetics of AS03 constituents in the mouse were consistent with AS03 inducing this pattern of inflammation. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-27T07:18:21.001473-05:
      DOI: 10.1002/jat.3130
  • Toxicity induced by Basic Violet 14, Direct Red 28 and Acid Red 26 in
           zebrafish larvae
    • Authors: Bing Shen; Hong‐Cui Liu, Wen‐Bin Ou, Grant Eilers, Sheng‐Mei Zhou, Fan‐Guo Meng, Chun‐Qi Li, Yong‐Quan Li
      Abstract: Basic Violet 14, Direct Red 28 and Acid Red 26 are classified as carcinogenic dyes in the European textile ecology standard, despite insufficient toxicity data. In this study, the toxicity of these dyes was assessed in a zebrafish model, and the underlying toxic mechanisms were investigated. Basic Violet 14 and Direct Red 28 showed acute toxicity with a LC50 value at 60.63 and 476.84 µg ml–1, respectively, whereas the LC50 of Acid Red 26 was between 2500 and 2800 µg ml–1. Treatment with Basic Violet 14, Direct Red 28 and Acid Red 26 resulted in common developmental abnormalities including delayed yolk sac absorption and swimming bladder deflation. Hepatotoxicity was observed in zebrafish treated with Basic Violet 14, and cardiovascular toxicity was found in zebrafish treated with Acid Red 26 at concentrations higher than 2500 µg ml–1. Basic Violet 14 also caused significant up‐regulation of GCLC gene expression in a dose‐dependent manner whereas Acid Red 26 induced significant up‐regulation of NKX2.5 and down‐regulation of GATA4 at a high concentration in a dose‐dependent manner. These results suggest that Basic Violet 14, Direct Red 28 and Acid Red 26 induce developmental and organ‐specific toxicity, and oxidative stress may play a role in the hepatotoxicity of Basic Violet 14, the suppressed GATA4 expression may have a relation to the cardiovascular toxicity of Acid Red 26. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-02-27T06:35:48.327256-05:
      DOI: 10.1002/jat.3134
  • Distribution and biomarkers of carbon‐14‐labeled fullerene C60
           ([14C(U)]C60) in female rats and mice for up to 30 days after intravenous
    • Authors: Susan C. J. Sumner; Rodney W. Snyder, Christopher Wingard, Ninell P. Mortensen, Nathan A. Holland, Jonathan H. Shannahan, Suraj Dhungana, Wimal Pathmasiri, Li Han, Anita H. Lewin, Timothy R. Fennell
      Abstract: A comprehensive distribution study was conducted in female rats and mice exposed to a suspension of uniformly carbon‐14‐labeled C60 ([14C(U)]C60). Rodents were administered [14C(U)]C60 (~0.9 mg kg−1 body weight) or 5% polyvinylpyrrolidone‐saline vehicle alone via a single tail vein injection. Tissues were collected at 1 h and 1, 7, 14 and 30 days after administration. A separate group of rodents received five daily injections of suspensions of either [14C(U)]C60 or vehicle with tissue collection 14 days post exposure. Radioactivity was detected in over 20 tissues at all time points. The highest concentration of radioactivity in rodents at each time point was in liver, lungs and spleen. Elimination of [14C(U)]C60 was < 2% in urine and feces at any 24 h time points. [14C(U)]C60 and [14C(U)]C60‐retinol were detected in liver of rats and together accounted for ~99% and ~56% of the total recovered at 1 and 30 days postexposure, respectively. The blood radioactivity at 1 h after [14C(U)]C60 exposure was fourfold higher in rats than in mice; blood radioactivity was still in circulation at 30 days post [14C(U)]C60 exposure in both species (
      PubDate: 2015-02-27T06:04:19.725928-05:
      DOI: 10.1002/jat.3110
  • Sertoli cell as a model in male reproductive toxicology: Advantages and
    • Authors: Mariana M. S. Reis; Ana C. Moreira, Mário Sousa, Premendu P. Mathur, Pedro F. Oliveira, Marco G. Alves
      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.
      PubDate: 2015-02-18T22:01:36.896633-05:
      DOI: 10.1002/jat.3122
  • The effect of a methyl‐deficient diet on the global DNA methylation
           and the DNA methylation regulatory pathways
    • Authors: Shota Takumi; Kazuyuki Okamura, Hiroyuki Yanagisawa, Tomoharu Sano, Yayoi Kobayashi, Keiko Nohara
      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.
      PubDate: 2015-02-17T19:31:53.091825-05:
      DOI: 10.1002/jat.3117
  • Heterozygous p53 knockout mouse model for dehydropyrrolizidine
           alkaloid‐induced carcinogenesis
    • Authors: Ammon W. Brown; Bryan L. Stegelmeier, Steven M. Colegate, Kip E. Panter, Edward L. Knoppel, Jeffery O. Hall
      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.
      PubDate: 2015-02-17T19:17:03.837317-05:
      DOI: 10.1002/jat.3120
  • Involvement of mitogen‐activated protein kinase and NF‐κB
           signaling pathways in perfluorooctane sulfonic acid‐induced
           inflammatory reaction in BV2 microglial cells
    • Authors: Jingying Zhu; Wenyi Qian, Yixin Wang, Rong Gao, Jun Wang, Hang Xiao
      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.
      PubDate: 2015-02-12T07:33:11.644658-05:
      DOI: 10.1002/jat.3119
  • 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
      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
      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
      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 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
      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
  • 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
      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
  • 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
  • T‐helper cell‐mediated factors in drug‐induced liver
    • Authors: Xinzhi Wang; Luyong Zhang, Zhenzhou Jiang
      First page: 695
      Abstract: Drug‐induced liver injury (DILI) leads to a large burden on the healthcare system due to its potential morbidity and mortality. The key for predicting and preventing DILI is to understand the underlying mechanisms. Hepatic inflammation is one of the most common features of DILI. The inflammation can be attributed to the innate immune response. The adaptive immune system is also affected by the innate immune response resulting in liver damage. T‐helper cells are important regulators of acquired immunity. T‐helper cell‐mediated immune responses play pivotal roles in the pathogenesis of a variety of liver disorders. This review summarizes recent advances in the T‐helper cell‐mediated factors in DILI and potential mechanisms, which may lead to a better understanding of DILI. Copyright © 2015 John Wiley & Sons, Ltd.
      PubDate: 2015-03-06T08:01:17.043191-05:
      DOI: 10.1002/jat.3115
  • 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
      Pages: 701 - 708
      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
  • Reactive oxygen species‐dependent JNK downregulated
           olaquindox‐induced autophagy in HepG2 cells
    • Authors: Dongxu Zhao; Congcong Wang, Shusheng Tang, Chaoming Zhang, Shen Zhang, Yan Zhou, Xilong Xiao
      Pages: 709 - 716
      Abstract: Autophagy plays an important role in response to intracellular and extracellular stress to sustain cell survival. However, dysregulated or excessive autophagy may lead to cell death, known as “type II programmed cell death,” and it is closely associated with apoptosis. In our previous study, we proposed that olaquindox induced apoptosis of HepG2 cells through a caspase‐9 dependent mitochondrial pathway. In this study, we investigated autophagy induced by olaquindox and explored the crosstalk between apoptosis and autophagy in olaquindox‐treated HepG2 cells. Olaquindox‐induced autophagy was demonstrated by the accumulation of monodansylcadervarine, as well as elevated expression of autophagy‐related MAP‐LC3 and Beclin 1 proteins. The autophagy inhibitor 3‐methyladenine significantly increased the apoptotic rate induced by olaquindox, which was correlated with increased ratio of Bax/Bcl‐2. The further studies showed that olaquindox increased the levels of reactive oxygen species (ROS), and antioxidant N‐acetyl‐L‐cysteine (NAC) effectively blocked the accumulation of ROS but failed to block autophagy. Moreover, olaquindox induced the activation of c‐Jun N‐terminal protein kinase (JNK), and JNK inhibitor SP600125 failed to block autophagy. Instead, olaquindox‐induced autophagy was enhanced by NAC or SP600125. Meanwhile, JNK activation was remarkably blocked by NAC, indicating that ROS may be the upstream signaling molecules of JNK activation and involved in the negative regulation of olaquindox‐induced autophagy. These results suggest that olaquindox induces autophagy in HepG2 cells and that olaquindox‐induced apoptosis can be enhanced by 3‐methyladenine. Olaquindox‐induced autophagy in HepG2 cells is upregulated by Beclin 1 but downregulated by ROS‐dependent JNK. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-18T04:30:06.618358-05:
      DOI: 10.1002/jat.3022
  • 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
      Pages: 717 - 728
      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
  • The relationship between chemical‐induced kidney weight increases
           and kidney histopathology in rats
    • Authors: Evisabel A. Craig; Zhongyu Yan, Q. Jay Zhao
      Pages: 729 - 736
      Abstract: The kidney is a major site of chemical excretion, which results in its propensity to exhibit chemically‐induced toxicological effects at a higher rate than most other organs. Although the kidneys are often weighed in animal toxicity studies, the manner in which these kidney weight measurements are interpreted and the value of this information in predicting renal damage remains controversial. In this study we sought to determine whether a relationship exists between chemically‐induced kidney weight changes and renal histopathological alterations. We also examined the relative utility of absolute and relative (kidney‐to‐body weight ratio) kidney weight in the prediction of renal toxicity. For this, data extracted from oral chemical exposure studies in rats performed by the National Toxicology Program were qualitatively and quantitatively evaluated. Our analysis showed a statistically significant correlation between absolute, but not relative, kidney weight and renal histopathology in chemically‐treated rats. This positive correlation between absolute kidney weight and histopathology was observed even with compounds that statistically decreased terminal body weight. Also, changes in absolute kidney weight, which occurred at subchronic exposures, were able to predict the presence or absence of kidney histopathology at both subchronic and chronic exposures. Furthermore, most increases in absolute kidney weight reaching statistical significance (irrespective of the magnitude of change) were found to be relevant for the prediction of histopathological changes. Hence, our findings demonstrate that the evaluation of absolute kidney weight is a useful method for identifying potential renal toxicants. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:42:22.00032-05:0
      DOI: 10.1002/jat.3036
  • 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
      Pages: 737 - 751
      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
  • 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: 752 - 758
      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
  • Early chronic lead exposure reduces exploratory activity in young C57BL/6J
    • Authors: Mayra Gisel Flores‐Montoya; Christina Sobin
      Pages: 759 - 765
      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
  • 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
      Pages: 766 - 780
      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
  • Comparison of the kinetics of various biomarkers of benzo[a]pyrene
           exposure following different routes of entry in rats
    • Authors: Marjory Moreau; Michèle Bouchard
      Pages: 781 - 790
      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
  • 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
      Pages: 791 - 798
      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
  • Assessment of temperature‐induced hERG channel blockade variation by
    • Authors: Rahul R. Kauthale; Shruta S. Dadarkar, Raghib Husain, Vikas V. Karande, Madhumanjiri M. Gatne
      Pages: 799 - 805
      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
  • 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
      Pages: 806 - 816
      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
  • 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
      Pages: 817 - 823
      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
  • Acute toxicity of 50 metals to Daphnia magna
    • Authors: Akira Okamoto; Masumi Yamamuro, Norihisa Tatarazako
      Pages: 824 - 830
      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
  • 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
      Pages: 831 - 841
      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
      Pages: 842 - 850
      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
  • 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
      Pages: 851 - 860
      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
  • 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
      Pages: 861 - 869
      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
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