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  Subjects -> ENVIRONMENTAL STUDIES (Total: 767 journals)
    - ENVIRONMENTAL STUDIES (704 journals)
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ENVIRONMENTAL STUDIES (704 journals)            First | 1 2 3 4 5 6 7 8     

International Innovation - climate     Open Access  
International innovation. Environment     Open Access  
International Journal of Acarology     Hybrid Journal   (Followers: 1)
International Journal of Advancement in Earth and Enviromental Sciences     Open Access   (Followers: 2)
International Journal of African Renaissance Studies - Multi-, Inter- and Transdisciplinarity     Hybrid Journal   (Followers: 2)
International Journal of Agricultural and Environmental Information Systems     Full-text available via subscription   (Followers: 1)
International Journal of Alternative Propulsion     Hybrid Journal   (Followers: 1)
International Journal of Applied Psychoanalytic Studies     Hybrid Journal   (Followers: 2)
International Journal of Chinese Culture and Management     Hybrid Journal   (Followers: 1)
International Journal of Corrosion     Open Access   (Followers: 10)
International Journal of Critical Infrastructures     Hybrid Journal   (Followers: 3)
International Journal of Disaster Risk Reduction     Hybrid Journal   (Followers: 5)
International Journal of Disaster Risk Science     Open Access   (Followers: 8)
International Journal of Ecological Economics and Statistics     Full-text available via subscription   (Followers: 1)
International Journal of Ecology     Open Access   (Followers: 8)
International Journal of Ecology & Development     Full-text available via subscription   (Followers: 1)
International Journal of Energy and Environmental Engineering     Open Access   (Followers: 2)
International Journal of Environment     Open Access   (Followers: 3)
International Journal of Environment and Health     Hybrid Journal   (Followers: 7)
International Journal of Environment and Pollution     Hybrid Journal   (Followers: 6)
International Journal of Environment and Sustainable Development     Hybrid Journal   (Followers: 14)
International Journal of Environment and Waste Management     Hybrid Journal   (Followers: 5)
International Journal of Environment, Workplace and Employment     Hybrid Journal   (Followers: 3)
International Journal of Environmental Engineering     Hybrid Journal   (Followers: 5)
International Journal of Environmental Health Engineering     Open Access  
International Journal of Environmental Health Research     Hybrid Journal   (Followers: 3)
International Journal of Environmental Policy and Decision Making     Hybrid Journal   (Followers: 10)
International Journal of Environmental Protection     Open Access   (Followers: 12)
International Journal of Environmental Research and Public Health     Open Access   (Followers: 14)
International Journal of Environmental Science and Technology     Hybrid Journal   (Followers: 5)
International Journal of Environmental Studies     Hybrid Journal   (Followers: 11)
International Journal of Exergy     Hybrid Journal   (Followers: 3)
International Journal of Forest, Soil and Erosion     Open Access   (Followers: 3)
International Journal of Global Environmental Issues     Hybrid Journal   (Followers: 4)
International Journal of Global Warming     Hybrid Journal   (Followers: 5)
International Journal of Greenhouse Gas Control     Partially Free   (Followers: 6)
International Journal of Health Planning and Management     Hybrid Journal   (Followers: 6)
International Journal of Hygiene and Environmental Health     Hybrid Journal   (Followers: 6)
International Journal of Logistics Research and Applications : A Leading Journal of Supply Chain Management     Hybrid Journal   (Followers: 9)
International Journal of Philosophical Studies     Hybrid Journal   (Followers: 2)
International Journal of Phytoremediation     Hybrid Journal   (Followers: 1)
International Journal of Process Systems Engineering     Hybrid Journal   (Followers: 1)
International Journal of Recycling of Organic Waste in Agriculture     Open Access   (Followers: 1)
International Journal of Regulation and Governance     Hybrid Journal   (Followers: 2)
International Journal of Reliability and Safety     Hybrid Journal   (Followers: 5)
International Journal of Renewable Energy Development     Open Access   (Followers: 5)
International Journal of Social Sciences and Management     Open Access  
International Journal of Soil, Sediment and Water     Open Access   (Followers: 8)
International Journal of Stress Management     Full-text available via subscription   (Followers: 8)
International Journal of Sustainable Construction Engineering and Technology     Open Access   (Followers: 7)
International Journal of Sustainable Engineering     Hybrid Journal   (Followers: 7)
International Journal of Sustainable Materials and Structural Systems     Hybrid Journal   (Followers: 5)
International Journal of Sustainable Society     Hybrid Journal   (Followers: 7)
International Journal of Testing     Hybrid Journal   (Followers: 1)
International Journal of the Commons     Open Access   (Followers: 2)
International Journal of Toxicology     Hybrid Journal   (Followers: 6)
International Journal of Water Resources and Environmental Engineering     Open Access   (Followers: 1)
International Review of Environmental and Resource Economics     Full-text available via subscription  
International Studies in the Philosophy of Science     Hybrid Journal   (Followers: 9)
Interventions : International Journal of Postcolonial Studies     Hybrid Journal   (Followers: 5)
IOP Conference Series: Earth and Environmental Science     Open Access   (Followers: 7)
Iranian Journal of Environmental Health Science & Engineering     Open Access   (Followers: 1)
Iranian Studies     Hybrid Journal   (Followers: 7)
Irish Educational Studies     Hybrid Journal   (Followers: 1)
Irish Journal of Earth Sciences     Full-text available via subscription  
Irish Political Studies     Hybrid Journal   (Followers: 7)
ISLE: Interdisciplinary Studies in Literature and Environment     Hybrid Journal   (Followers: 2)
Isotopes in Environmental and Health Studies     Hybrid Journal   (Followers: 2)
Israel Studies     Full-text available via subscription   (Followers: 5)
ISRN Ecology     Open Access  
ISRN Environmental Chemistry     Open Access  
Jahangirnagar University Environmental Bulletin     Open Access  
Journal of Bioremediation & Biodegradation     Open Access   (Followers: 2)
Journal of Earth Science & Climatic Change     Open Access   (Followers: 8)
Journal of Petroleum & Environmental Biotechnology     Open Access   (Followers: 2)
Journal of Advances in Environmental Health Research     Open Access  
Journal of Agricultural and Environmental Ethics     Hybrid Journal   (Followers: 6)
Journal of Agricultural Biotechnology and Sustainable Development     Open Access  
Journal of Agriculture and Environment     Open Access  
Journal of Agriculture and Environment for International Development     Open Access   (Followers: 6)
Journal of Agrobiology     Open Access   (Followers: 2)
Journal of Applied Ecology     Hybrid Journal   (Followers: 156)
Journal of Applied Meteorology and Climatology     Full-text available via subscription   (Followers: 7)
Journal of Applied Psychoanalytic Studies     Hybrid Journal   (Followers: 1)
Journal of Applied Sciences and Environmental Management     Open Access   (Followers: 1)
Journal of Applied Sciences in Environmental Sanitation     Open Access   (Followers: 6)
Journal of Applied Toxicology     Hybrid Journal   (Followers: 9)
Journal of Applied Volcanology     Open Access   (Followers: 6)
Journal of Arid Environments     Hybrid Journal   (Followers: 8)
Journal of Asian Studies     Full-text available via subscription   (Followers: 21)
Journal of Biochemical and Molecular Toxicology     Hybrid Journal   (Followers: 5)
Journal of Black Studies     Hybrid Journal   (Followers: 2)
Journal of Chemical Ecology     Hybrid Journal   (Followers: 1)
Journal of Chemical Health and Safety     Hybrid Journal   (Followers: 2)
Journal of Climate     Full-text available via subscription   (Followers: 23)
Journal of Coastal Research     Full-text available via subscription   (Followers: 11)
Journal of Contaminant Hydrology     Hybrid Journal   (Followers: 9)
Journal of Contemporary European Studies     Hybrid Journal   (Followers: 4)
Journal of East African Natural History     Full-text available via subscription   (Followers: 5)
Journal of Ecology     Hybrid Journal   (Followers: 35)

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Journal Cover Journal of Applied Toxicology
   Journal TOC RSS feeds Export to Zotero [11 followers]  Follow    
   Hybrid Journal Hybrid journal (It can contain Open Access articles)
     ISSN (Print) 0260-437X - ISSN (Online) 1099-1263
     Published by John Wiley and Sons Homepage  [1603 journals]   [SJR: 0.689]   [H-I: 47]
  • Diagnostic and predictive performance and standardized threshold of
           traditional biomarkers for drug‐induced liver injury in rats
    • Authors: Yutaka Tonomura; Yuki Kato, Hiroyuki Hanafusa, Yuji Morikawa, Keigo Matsuyama, Takeki Uehara, Motonobu Ueno, Mikinori Torii
      Pages: n/a - n/a
      Abstract: Traditional biomarkers such as alanine and aspartate aminotransferase (ALT, AST) and total bilirubin (TBIL) have been widely used for detecting drug‐induced liver injury (DILI). Although the Food and Drug Administration (FDA) proposed standardized thresholds for human as Hy's law, those for animals have not been determined, and predictability of these biomarkers for future onset of hepatic lesions remains unclear. In this study, we investigated these diagnostic and predictive performance of 10 traditional biomarkers for liver injury by receiver‐operating characteristic (ROC) curve, using a free‐access database where 142 hepatotoxic or non‐hepatotoxic compounds were administrated to male rats (n = 5253). Standardization of each biomarker value was achieved by calculating the ratio to control mean value, and the thresholds were determined under the condition of permitting 5% false positive. Of these 10 biomarkers, AST showed the best diagnostic performance. Furthermore, ALT and TBIL also showed high performance under the situation of hepatocellular necrosis and bile duct injury, respectively. Additionally, the availability of the diagnostic thresholds in difference testing facility was confirmed by the application of these thresholds to in‐house prepared dataset. Meanwhile, incorrect diagnosis by the thresholds was also observed. Regarding prediction, all 10 biomarkers showed insufficient performance for future onset of hepatic lesions. In conclusion, the standardized diagnostic thresholds enable consistent evaluation of traditional biomarkers among different facilities, whereas it was suggested that novel biomarker is required for more accurate diagnosis and prediction of DILI. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-04T05:02:30.374456-05:
      DOI: 10.1002/jat.3053
  • Perturbation of cytosolic calcium by 2‐aminoethoxydiphenyl borate
           and caffeine affects zebrafish myofibril alignment
    • Authors: Hsin‐Ju Wu; Tsorng‐Harn Fong, Shen‐Liang Chen, Jen‐Cheng Wei, I‐Jong Wang, Chi‐Chung Wen, Chao‐Yuan Chang, Xing‐Guang Chen, Wei‐Yu Chen, Hui‐Min Chen, Juin‐Lin Horng, Yun‐Hsin Wang, Yau‐Hung Chen
      Pages: n/a - n/a
      Abstract: The objective of the current study was to investigate the effects of Ca2+ levels on myofibril alignment during zebrafish embryogenesis. To investigate how altered cytoplasmic Ca2+ levels affect myofibril alignment, we exposed zebrafish embryos to 2‐aminothoxyldiphenyl borate (2‐APB; an inositol 1,4,5‐trisphosphate receptor inhibitor that reduces cytosolic Ca2+ levels) and caffeine (a ryanodine receptor activator that enhances cytosolic Ca2+ levels). The results demonstrated that the most evident changes in zebrafish embryos treated with 2‐APB were shorter body length, curved trunk and malformed somite boundary. In contrast, such malformed phenotypes were evident neither in untreated controls nor in caffeine‐treated embryos. Subtle morphological changes, including changes in muscle fibers, F‐actin and ultrastructures were easily observed by staining with specific monoclonal antibodies (F59 and α‐laminin), fluorescent probes (phalloidin) and by transmission electron microscopy. Our data suggested that: (1) the exposure to 2‐APB and/or caffeine led to myofibril misalignment; (2) 2‐APB‐treated embryos displayed split and short myofibril phenotypes, whereas muscle fibers from caffeine‐treated embryos were twisted and wavy; and (3) zebrafish embryos co‐exposed to 2‐APB and caffeine resulted in normal myofibril alignment. In conclusion, we proposed that cytosolic Ca2+ is important for myogenesis, particularly for myofibril alignment. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-04T04:56:51.010578-05:
      DOI: 10.1002/jat.3057
  • Skin absorption of six performance amines used in metalworking fluids
    • Authors: Lauriane N. Roux; James D. Brooks, James L. Yeatts, Ronald E. Baynes
      Pages: n/a - n/a
      Abstract: Every year, 10 million workers are exposed to metalworking fluids (MWFs) that may be toxic. There are four types of MWFs: neat oils and three water‐based MWFs (soluble oil, semisynthetic and synthetic), which are diluted with water and whose composition varies according to the mineral oils ratio. MWFs also contain various additives. To determine the absorption of six amines used as corrosion inhibitors and biocides in MWFs, porcine skin flow‐through diffusion cell experiments were conducted with hydrophilic ethanolamines (mono‐, di‐ and triethanolamine, MEA, DEA and TEA respectively) and a mixture of lipophilic amines (dibutylethanolamine, dicyclohexylamine and diphenylamine). The six amines were dosed in four vehicles (water and three generic water‐based MWF formulations) and analyzed using a scintillation counter or gas chromatography/mass spectrometry. These 24 h studies showed that dermal absorption significantly (P 
      PubDate: 2014-09-04T04:56:30.081537-05:
      DOI: 10.1002/jat.3056
  • A representative retinoid X receptor antagonist UVI3003 induced
           teratogenesis in zebrafish embryos
    • Authors: Liang Zheng; Ting Xu, Daoji Li, Junliang Zhou
      Pages: n/a - n/a
      Abstract: Retinoid X receptor (RXR) interfering activity has been detected in different water resources. To study RXR disruptor‐induced toxicological effects on vertebrates, embryos of zebrafish (Danio rerio) were exposed to a representative RXR antagonist UVI3003. Results showed that the teratogenic index (LC50/EC50) of UVI3003 was as high as 5.4. UVI3003 induced multiple malformations of embryos, including deformed fins, reduced brains, small jaws, bent tails and edema in hearts, the degree of which became more severe with increasing exposure concentration. Although no significant difference was observed in the hatching rates between the exposure group and control, the whole body length was significantly reduced by 6.5% and 8.9% when exposed to 200 and 300 µg l−1 of UVI3003, respectively. The heart rate also significantly decreased by 8.8–50.2% during exposure. Further experiments revealed that the pharyngula stage was the most sensitive development phase in terms of embryo response to UVI3003. The results demonstrated severe teratogenicity of RXR antagonist in zebrafish embryos and provided important data for ecotoxicological evaluation of RXR antagonists. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-03T03:07:21.862962-05:
      DOI: 10.1002/jat.3051
  • Immunophenotyping does not improve predictivity of the local lymph node
           assay in mice
    • Authors: Volker Strauss; Susanne N. Kolle, Naveed Honarvar, Martina Dammann, Sibylle Groeters, Frank Faulhammer, Robert Landsiedel, Bennard Ravenzwaay
      Pages: n/a - n/a
      Abstract: The local lymph node assay (LLNA) is a regulatory accepted test for the identification of skin sensitizing substances by measuring radioactive thymidine incorporation into the lymph node. However, there is evidence that LLNA is overestimating the sensitization potential of certain substance classes in particular those exerting skin irritation. Some reports describe the additional use of flow cytometry‐based immunophenotyping to better discriminate irritants from sensitizing irritants in LLNA. In the present study, the 22 performance standards plus 8 surfactants were assessed using the radioactive LLNA method. In addition, lymph node cells were immunophenotyped to evaluate the specificity of the lymph node response using cell surface markers such as B220 or CD19, CD3, CD4, CD8, I‐Aκ and CD69 with the aim to allow a better discrimination above all between irritants and sensitizers, but also non‐irritating sensitizers and non‐sensitizers. However, the markers assessed in this study do not sufficiently differentiate between irritants and irritant sensitizers and therefore did not improve the predictive capacity of the LLNA. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-03T03:06:12.16689-05:0
      DOI: 10.1002/jat.3042
  • Reversible cholinesterase inhibitors as pre‐treatment for exposure
           to organophosphates: assessment using azinphos‐methyl
    • Authors: Georg A. Petroianu; Syed M. Nurulain, Mohamed Y. Hasan, Kamil Kuča, Dietrich E. Lorke
      Pages: n/a - n/a
      Abstract: Pre‐treatment with reversible acetylcholinesterase (AChE) inhibitors before organophosphorous compound (OPC) exposure can reduce OPC‐induced mortality. However, pyridostigmine, the only substance employed for such prophylaxis, is merely efficacious against a limited number of OPCs. In search of more efficacious and broad‐range alternatives, we have compared in vivo the ability of five reversible AChE inhibitors (pyridostigmine, physostigmine, ranitidine, tacrine and K‐27) to reduce mortality induced by the OPC azinphos‐methyl. Protection was quantified using Cox analysis by determining the relative risk (RR) of death in rats that were administered these AChE inhibitors in equitoxic dosage (25% of LD01) 30 min before azinphos‐methyl exposure. Azinphos‐methyl‐induced mortality was significantly reduced by all five tested compounds as compared with the reference group that was only exposed to azinphos‐methyl without prior pre‐treatment (RR = 1). The most efficacious prophylactic agents were K‐27 (RR = 0.15) and physostigmine (RR = 0.21), being significantly more efficacious than ranitidine (RR = 0.62) and pyridostigmine (RR = 0.37). Pre‐treatment with tacrine (RR = 0.29) was significantly more efficacious than pre‐treatment with ranitidine, but the difference between tacrine and pyridostigmine was not significant. Our results indicate that prophylactic administration of the oxime K‐27 may be a promising alternative in cases of imminent OPC exposure. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-03T03:04:57.769676-05:
      DOI: 10.1002/jat.3052
  • Carnosic acid induces autophagic cell death through inhibition of the
           Akt/mTOR pathway in human hepatoma cells
    • Authors: Qilong Gao; Huaimin Liu, Yamin Yao, Liang Geng, Xinfeng Zhang, Lifeng Jiang, Bian Shi, Feng Yang
      Abstract: The therapeutic goal of cancer treatment is now geared towards triggering tumour‐selective cell death with autophagic cell death being required for the chemotherapy of apoptosis‐resistant cancer. In this study, Carnosic acid (CA), a polyphenolic diterpene isolated from Rosemary (Rosemarinus officinalis), significantly induced autophagic cell death in HepG2 cells. Ca treatment caused the formation of autophagic vacuoles produced an increasing ratio of LC3‐II to LC3‐I in a time‐ and dose‐dependent manner but had no effect on the levels of autophagy‐related protein ATG6 and ATG13 expression. Autophagy inhibitors, 3‐methyladenine (3‐MA), chloroquine and bafilomycin A1, or ATG genes silencing in HepG2 cells significantly inhibited CA‐induced autophagic cell death. The CA treatment decreased the levels of phosphorylated Akt and mTOR without any effects on PI3K or PTEN. Most importantly, overexpression of Akt and knockdown of PTEN attenuated autophagy induction in CA‐treated cells. Taken together, our results indicated that CA induced autophagic cell death through inhibition of the Akt/mTOR pathway in human hepatoma cells. These findings suggest that CA has a great potential for the treatment of hepatoma via autophagic induction. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-01T07:01:21.780556-05:
      DOI: 10.1002/jat.3049
  • Hydrophobic sodium fluoride‐based nanocrystals doped with lanthanide
           ions: assessment of in vitro toxicity to human blood lymphocytes and
    • Authors: Bartlomiej Sojka; Miroslava Kuricova, Aurelia Liskova, Maria Bartusova, Mateusz Banski, Jan Misiewicz, Maria Dusinska, Mira Horvathova, Eva Jahnova, Silvia Ilavska, Michaela Szabova, Eva Rollerova, Artur Podhorodecki, Jana Tulinska
      Abstract: In vitro immunotoxicity of hydrophobic sodium fluoride‐based nanocrystals (NCs) doped with lanthanide ions was examined in this study. Although there is already a significant amount of optical and structural data on NaYF4 NCs, data on safety assessment are missing. Therefore, peripheral whole blood from human volunteers was used to evaluate the effect of 25 and 30 nm hydrophobic NaYF4 NCs dissolved in cyclohexane (CH) on lymphocytes, and of 10 nm NaYF4 NCs on phagocytes. In the concentration range 0.12–75 µg cm−2 (0.17–106 µg ml−1), both 25 and 30nm NaYF4 NCs did not induce cytotoxicity when measured as incorporation of [3H]‐thymidine into DNA. Assessment of lymphocyte function showed significant suppression of the proliferative activity of T‐lymphocytes and T‐dependent B‐cell response in peripheral blood cultures (n = 7) stimulated in vitro with mitogens phytohemagglutinin (PHA) and pokeweed (PWM) (PHA > PWM). No clear dose–response effect was observed. Phagocytic activity and respiratory burst of leukocytes (n = 5–8) were generally less affected. A dose‐dependent suppression of phagocytic activity of granulocytes in cultures treated with 25 nm NCs was observed (vs. medium control). A decrease in phagocytic activity of monocytes was found in cells exposed to higher doses of 10 and 30 nm NCs. The respiratory burst of phagocytes was significantly decreased by exposure to the middle dose of 30 nm NCs only. In conclusion, our results demonstrate immunotoxic effects of hydrophobic NaYF4 NCs doped with lanthanide ions to lymphocytes and to lesser extent to phagocytes. Further research needs to be done, particularly faze transfer of hydrophobic NCs to hydrophilic ones, to eliminate the solvent effect. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-09-01T07:01:07.943675-05:
      DOI: 10.1002/jat.3050
  • Protective effects of ascorbic acid against the genetic and epigenetic
           alterations induced by 3,5‐dimethylaminophenol in AA8 cells
    • Authors: Ming‐Wei Chao; P𝚤nar Erkekoglu, Chia‐Yi Tseng, Wenjie Ye, Laura J. Trudel, Paul L. Skipper, Steven R. Tannenbaum, Gerald N. Wogan
      Abstract: Exposure to monocyclic aromatic alkylanilines (MAAs), namely 2,6‐dimethylaniline (2,6‐DMA), 3,5‐dimethylaniline (3,5‐DMA) and 3‐ethylaniline (3‐EA), was significantly and independently associated with bladder cancer incidence. 3,5‐DMAP (3,5‐dimethylaminophenol), a metabolite of 3,5‐DMA, was shown to induce an imbalance in cytotoxicity cellular antioxidant/oxidant status, and DNA damage in mammalian cell lines. This study was designed to evaluate the protective effect of ascorbic acid (Asc) against the cytotoxicity, reactive oxygen species (ROS) production, genotoxicity and epigenetic changes induced by 3,5‐DMAP in AA8 Chinese Hamster Ovary (CHO) cells. In different cellular fractions, 3,5‐DMAP caused alterations in the enzyme activities orchestrating a cellular antioxidant balance, decreases in reduced glutathione levels and a cellular redox ratio as well as increases in lipid peroxidation and protein oxidation. We also suggest that the cellular stress caused by this particular alkylaniline leads to both genetic (Aprt mutagenesis) and epigenetic changes in histones 3 and 4 (H3 and H4). This may further cause molecular events triggering different pathological conditions and eventually cancer. In both cytoplasm and nucleus, Asc provided increases in 3,5‐DMAP‐reduced glutathione levels and cellular redox ratio and decreases in the lipid peroxidation and protein oxidation. Asc was also found to be protective against the genotoxic and epigenetic effects initiated by 3,5‐DMAP. In addition, Asc supplied protection against the cell cycle (G1 phase) arrest induced by this particular alkylaniline metabolite.
      PubDate: 2014-09-01T06:51:00.294209-05:
      DOI: 10.1002/jat.3046
  • In utero and early childhood exposure to arsenic decreases lung function
           in children
    • Authors: Rogelio Recio‐Vega; Tania Gonzalez‐Cortes, Edgar Olivas‐Calderon, R. Clark Lantz, A. Jay Gandolfi, Cesar Gonzalez‐De Alba
      Abstract: The lung is a target organ for adverse health outcomes following exposure to As. Several studies have reported a high prevalence of respiratory symptoms and diseases in subjects highly exposed to As through drinking water; however, most studies to date has been performed in exposed adults, with little information on respiratory effects in children. The objective of the study was to evaluate the association between urinary levels of As and its metabolites with lung function in children exposed in utero and in early childhood to high As levels through drinking water. A total of 358 healthy children were included in our study. Individual exposure was assessed based on urinary concentration of inorganic As. Lung function was assessed by spirometry. Participants were exposed since pregnancy until early childhood to an average water As concentration of 152.13 µg l–1. The mean urinary As level registered in the studied subjects was 141.2 µg l–1 and only 16.7% had a urinary concentration below the national concern level. Forced vital capacity was significantly decreased in the studied population and it was negatively associated with the percentage of inorganic As. More than 57% of the subjects had a restrictive spirometric pattern. The urinary As level was higher in those children with restrictive lung patterns when compared with the levels registered in subjects with normal spirometric patterns. Exposure to As through drinking water during in utero and early life was associated with a decrease in forced vital capacity and with a restrictive spirometric pattern in the children evaluated. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-15T08:01:31.55848-05:0
      DOI: 10.1002/jat.3023
  • Development of haemostatic decontaminants for the treatment of wounds
           contaminated with chemical warfare agents. 1: Evaluation of in vitro
           clotting efficacy in the presence of certain contaminants
    • Authors: Charlotte A. Hall; Helen L. Lydon, Christopher H. Dalton, J. K. Chipman, John S. Graham, Robert P. Chilcott
      Abstract: The treatment of penetrating, haemorrhaging injuries sustained within a hazardous environment may be complicated by contamination with toxic chemicals. There are currently no specific medical countermeasures for such injuries. Haemostats with an absorbent mechanism of action have the potential to simultaneously stop bleeding and decontaminate wounds. However, a primary requirement of a ‘haemostatic decontaminant’ is the retention of clotting function in the presence of chemical contaminants. Thus, the aim of this study was to investigate the haemostatic efficacy of seven commercially available haemostats in the presence of toxic chemicals (soman, VX, sulphur mustard, petrol, aviation fuel and motor oil). Clot viscosity was assessed ex vivo using thrombelastography following treatment of pig blood with: (i) toxic chemical; (ii) haemostat; or (iii) haemostat in combination with toxic chemical. Several contaminants (VX, petrol and GD) were found to be pro‐haemostatic and none had an adverse effect on the rate with which the test products attained haemostasis. However, the total clot strength for blood treated with certain haemostats in the presence of sulphur mustard, soman and petrol was significantly decreased. Three test products failed to demonstrate haemostatic function in this ex vivo (thrombelastography) model; this was tentatively ascribed to the products achieving haemostasis through a tamponade mechanism of action, which can only be replicated using in vivo models. Overall, this study has identified a number of commercial products that may have potential as haemostatic decontaminants and warrant further investigation to establish their decontaminant efficacy. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-15T07:46:27.370339-05:
      DOI: 10.1002/jat.3019
  • Cardiolipin profiles as a potential biomarker of mitochondrial health in
           diet‐induced obese mice subjected to exercise,
           diet‐restriction and ephedrine treatment
    • Authors: Catherine Faber; Zhaohai J. Zhu, Stephen Castellino, David S. Wagner, Roger H. Brown, Richard A. Peterson, Lisa Gates, Joanna Barton, Mark Bickett, Laura Hagerty, Carie Kimbrough, Mario Sola, David Bailey, Holly Jordan, Chandikumar S. Elangbam
      Abstract: Cardiolipin (CL) is crucial for mitochondrial energy metabolism and structural integrity. Alterations in CL quantity or CL species have been associated with mitochondrial dysfunction in several pathological conditions and diseases, including mitochondrial dysfunction‐related compound attrition and post‐market withdrawal of promising drugs. Here we report alterations in the CL profiles in conjunction with morphology of soleus muscle (SM) and brown adipose tissue (BAT) in diet‐induced obese (DIO) mice, subjected to ephedrine treatment (EPH: 200 mg kg–1 day–1 orally), treadmill exercise (EX: 10 meters per min, 1 h per day), or dietary restriction (DR: 25% less of mean food consumed by the EX group) for 7 days. Mice from the DR and EPH groups had a significant decrease in percent body weight and reduced fat mass compared with DIO controls. Morphologic alterations in the BAT included brown adipocytes with reduced cytoplasmic lipid droplets and increased cytoplasmic eosinophilia in the EX, DR and EPH groups. Increased cytoplasmic eosinophilia in the BAT was ultrastructurally manifested by increased mitochondrial cristae, fenestration of mitochondrial cristae, increased electron density of mitochondrial matrix, and increased complexity of shape and elongation of mitochondria. Mitochondrial ultrastructural alterations in the SM of the EX and DR groups included increased mitochondrial cristae, cup‐shaped mitochondria and mitochondrial degeneration. All four CL species (tri‐linoleoyl‐mono‐docosahexaenoyl, tetralinoleoyl, tri‐linoleoyl‐mono‐oleoyl, and di‐linoleoyl‐di‐oleoyl) were increased in the BAT of the DR and EPH groups and in the SM of the EPH and EX groups. In conclusion, cardiolipin profiling supported standard methods for assessing mitochondrial biogenesis and health, and may serve as a potential marker of mitochondrial dysfunction in preclinical toxicity studies. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-15T07:38:38.650673-05:
      DOI: 10.1002/jat.3030
  • Systemic drugs inducing non‐immediate cutaneous adverse reactions
           and contact sensitizers evoke similar responses in THP‐1 cells
    • Authors: Margarida Gonçalo; João Martins, Ana Silva, Bruno Neves, Américo Figueiredo, Teresa Cruz, Celeste Lopes
      Abstract: Contact sensitizers induce phenotypic and functional changes in dendritic cells (DC) that enhance their antigen‐presenting capacity and, ultimately, modulate the T cell response. To evaluate if there is a similar effect of drugs causing T‐cell‐mediated cutaneous adverse drug reactions (CADR), we studied the in vitro effect of drugs on THP‐1 cells, a cell line widely used to evaluate the early molecular and cellular events triggered by contact sensitizers. The effect of allopurinol, oxypurinol, ampicillin, amoxicillin, carbamazepine and sodium valproate, at EC30 concentrations, was evaluated on p38 MAPK activation, by Western Blot, and on the expression of genes coding for DC maturation markers, pro‐inflammatory cytokine/chemokines and hemeoxygenase 1 (HMOX1), by real‐time RT‐PCR. Results were compared with lipopolysaccharide (LPS), a DC maturation stimulus, and the strong contact sensitizer, 1‐fluoro‐2,4‐dinitrobenzene (DNFB). All drugs studied significantly upregulated HMOX1 gene transcription and all, except the anticonvulsants, also upregulated IL8. Allopurinol and oxypurinol showed the most intense effect, in a magnitude similar to DNFB and superior to betalactams. Transcription of CD40, IL12B and CXCL10 genes by drugs was more irregular. Moreover, like DNFB, all drugs activated p38 MAPK, although significantly only for oxypurinol. Like contact sensitizers, drugs that cause non‐immediate CADR activate THP‐1 cells in vitro, using different signalling pathways and affecting gene transcription with an intensity that may reflect the frequency and severity of the CADR they cause. Direct activation of antigen‐presenting DC by systemic drugs may be an important early step in the pathophysiology of non‐immediate CADR. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:46:53.569007-05:
      DOI: 10.1002/jat.3033
  • Analysis of drugs of abuse in human plasma by dispersive
           liquid–liquid microextraction and high‐performance liquid
    • Authors: P. Fernández; M. Regenjo, A. M. Bermejo, A. M. Fernández, R. A. Lorenzo, A. M. Carro
      Abstract: Opioids and cocaine are widely used at present, both for recreational purposes and as drugs of abuse. This raises the need to develop new analytical methods specifically designed for the simultaneous detection of several drugs of abuse in biological samples. In this work, dispersive liquid–liquid microextraction (DLLME) was assessed as a new sample treatment for the simultaneous extraction of morphine (MOR), 6‐acetylmorphine (6AM), cocaine (COC), benzoylecgonine (BZE) and methadone (MET) from human plasma. Preliminary assays were done before developing an experimental design based on a Uniform Network Doehlert which allowed the optimum extraction conditions to be identified, namely: a volume of extractant solvent (chloroform) and dispersant solvent (acetonitrile) of 220 µl and 3.2 ml, respectively; 0.2 g of NaCl as a salting‐out additive; pH 10.6 and ultrasound stirring for 3.5 min. The resulting extracts were analyzed by high‐performance liquid chromatography with photodiode array detection (HPLC‐PDA), using an XBridge® RP18 column (250 × 4.6 mm i.d., 5 µm particle size). Calibration graphs were linear over the concentration range 0.1–10 µg ml–1, and detection limits ranged from 13.9 to 28.5 ng ml–1. Precision calculated at three different concentration levels in plasma was included in the range 0.1–6.8% RSD. Recoveries of the five drugs were all higher than 84% on average. Finally the proposed method was successfully applied to 22 plasma samples from heroin, cocaine and/or methadone users, and the most frequently detected drug was benzoylecgonine, followed by methadone, cocaine and morphine. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:42:27.742954-05:
      DOI: 10.1002/jat.3035
  • The relationship between chemical‐induced kidney weight increases
           and kidney histopathology in rats
    • Authors: Evisabel A. Craig; Zhongyu Yan, Q. Jay Zhao
      Abstract: The kidney is a major site of chemical excretion, which results in its propensity to exhibit chemically‐induced toxicological effects at a higher rate than most other organs. Although the kidneys are often weighed in animal toxicity studies, the manner in which these kidney weight measurements are interpreted and the value of this information in predicting renal damage remains controversial. In this study we sought to determine whether a relationship exists between chemically‐induced kidney weight changes and renal histopathological alterations. We also examined the relative utility of absolute and relative (kidney‐to‐body weight ratio) kidney weight in the prediction of renal toxicity. For this, data extracted from oral chemical exposure studies in rats performed by the National Toxicology Program were qualitatively and quantitatively evaluated. Our analysis showed a statistically significant correlation between absolute, but not relative, kidney weight and renal histopathology in chemically‐treated rats. This positive correlation between absolute kidney weight and histopathology was observed even with compounds that statistically decreased terminal body weight. Also, changes in absolute kidney weight, which occurred at subchronic exposures, were able to predict the presence or absence of kidney histopathology at both subchronic and chronic exposures. Furthermore, most increases in absolute kidney weight reaching statistical significance (irrespective of the magnitude of change) were found to be relevant for the prediction of histopathological changes. Hence, our findings demonstrate that the evaluation of absolute kidney weight is a useful method for identifying potential renal toxicants. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:42:22.00032-05:0
      DOI: 10.1002/jat.3036
  • Protective role of L‐ascorbic acid, N‐acetylcysteine and
           apocynin on neomycin‐induced hair cell loss in Zebrafish
    • Authors: Chia‐Yen Wu; Han‐Jung Lee, Chi‐Fang Liu, Mallikarjuna Korivi, Hwei‐Hsien Chen, Ming‐Huan Chan
      Abstract: Hair cells are highly sensitive to environmental insults and other therapeutic drugs. The adverse effects of drugs such as aminoglycosides can cause hair cell death and lead to hearing loss and imbalance. The objective of the present study was to evaluate the protective activity of L‐ascorbic acid, N‐acetylcysteine (NAC) and apocynin on neomycin‐induced hair cell damage in zebrafish (Danio rerio) larvae at 5 days post fertilization (dpf). Results showed that the loss of hair cells within the neuromasts of the lateral lines after neomycin exposure was evidenced by a significantly lower number of neuromasts labeled with fluorescent dye FM1‐43FX observed under a microscope. Co‐administration with L‐ascorbic acid, NAC and apocynin protected neomycin‐induced hair cell loss within the neuromasts. Moreover, these three compounds reduced the production of reactive oxygen species (ROS) in neuromasts exposed to neomycin, indicating that their antioxidant action is involved. In contrast, the neuromasts were labeled with specific fluorescent dye Texas‐red conjugated with neomycin to detect neomycin uptake. Interestingly, the uptake of neomycin into hair cells was not influenced by these three antioxidant compounds. These data imply that prevention of hair cell damage against neomycin by L‐ascorbic acid, NAC and apocynin might be associated with inhibition of excessive ROS production, but not related to modulating neomycin uptake. Our findings conclude that L‐ascorbic acid, NAC and apocynin could be used as therapeutic drugs to protect aminoglycoside‐induced listening impairment after further confirmatory studies. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:42:16.707002-05:
      DOI: 10.1002/jat.3043
  • Plasma miR‐208 as a useful biomarker for drug‐induced
           cardiotoxicity in rats
    • Authors: Yoko Nishimura; Chiaki Kondo, Yuji Morikawa, Yutaka Tonomura, Mikinori Torii, Jyoji Yamate, Takeki Uehara
      Abstract: Cardiotoxicity is one of the major safety concerns in drug development. Therefore, detecting and monitoring cardiotoxicity throughout preclinical and clinical studies is important for pharmaceutical companies. The present study was conducted in order to explore a plasma miRNA biomarker for cardiotoxicity in rats. As organ specificity is an important factor for a biomarker, we analyzed the miRNA microarray dataset in 55 organs/tissues in normal male rats. Based on this analysis, 5 miRNAs consisting of miR‐208 (heart‐specific), miR‐1, miR‐133a, miR‐133b (heart and skeletal muscle‐specific) and miR‐206 (skeletal muscle‐specific) were selected. Next, we evaluated the usefulness of those 5 miRNAs as circulating biomarkers in rats administered with single‐dose isoproterenol or doxorubicin. Plasma miR‐208 was consistently increased through 24 h after dosing in rats administered with isoproterenol, whereas plasma concentrations of cardiac troponin (cTn) showed transient elevation. In contrast, the plasma levels of miR‐1, miR‐133a, miR‐133a and miR‐206 were elevated after treatment with doxorubicin, probably as a result of skeletal muscle toxicity. Additionally, the plasma miR‐208 level was elevated even after repeat‐dose administration (once daily for 7 days) of isoproterenol under which the pathological condition proceeded to the sub‐chronic phase such as fibrosis. Thus, our data suggest that miR‐208 is a promising plasma biomarker for cardiotoxicity in rats. Monitoring of plasma miR‐208 levels in rats may lead to more accurate evaluation of cardiotoxicity in preclinical studies. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-08-04T09:03:05.796088-05:
      DOI: 10.1002/jat.3044
  • In vitro evaluation of the effects of perfluorooctanesulfonic acid (PFOS)
           and perfluorooctanoic acid (PFOA) on IL‐2 production in human
    • Authors: Kristin Midgett; Margie M. Peden‐Adams, Gary S. Gilkeson, Diane L. Kamen
      Abstract: Perfluorinated compounds, such as perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA), have been shown to alter various immune functions suggesting they are immunotoxic. This study assessed the effects of PFOS and PFOA on interleukin (IL)‐2 production in the human Jurkat T‐cell line and PFOS in healthy human primary T cells. Jurkat cells were stimulated with phytohemagglutinin (PHA)/phorbol myristate acetate (PMA), anti CD‐3/anti CD‐28, or anti CD‐3, and dosed with 0, 0.05, 0.1, 0.5, 1, 5, 10, 50, 75, or 100 µg ml−1 PFOS or 0, 0.005, 0.01, 0.05, 0.1, 0.5, 1, 5, or 10 µg ml−1 PFOA. Jurkat cells stimulated with PHA/PMA or anti CD‐3 exhibited decreased IL‐2 production beginning at 50 µg PFOS ml−1 and 5 µg PFOS ml−1 respectively, but stimulation with anti‐CD3/anti‐CD28 resulted in no changes compared with the control. Addition of the PPAR‐alpha antagonist GW6471 to PFOS‐dosed cells stimulated with PHA/PMA resulted in decreases in IL‐2 production starting at 50 µg PFOS ml−1, which suggests PFOS affected T‐cell IL‐2 production via PPAR‐alpha‐independent mechanisms. Exposure to PFOA, PFOA + GW6471, or PFOS + PFOA in Jurkat cells resulted in no significant differences in IL‐2 production. In vitro dosing studies using healthy primary human CD4+ T cells were consistent with the Jurkat results. These data demonstrated that PFOA did not impact IL‐2 production, but PFOS suppressed IL‐2 production in both a human cell line and human primary cells at dose levels within the high end of the human exposure range. A decrease in IL‐2 production is characteristic of autoimmune diseases such as systemic lupus erythematosus and should be further investigated. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-23T08:44:08.47006-05:0
      DOI: 10.1002/jat.3037
  • Characterization of Oryzias latipes glucocorticoid receptors and their
           unique response to progestins
    • Authors: Shinichi Miyagawa; Anke Lange, Saki Tohyama, Yukiko Ogino, Takeshi Mizutani, Tohru Kobayashi, Norihisa Tatarazako, Charles R. Tyler, Taisen Iguchi
      Abstract: Various receptor bioassays, including estrogens, androgens and thyroid hormones, have been developed and applied successfully for assessing hormone function in a wide range of animal species, including fish. In fish, corticosteroids play a pivotal role in physiology as they do in mammals, but far less is known about the corticosteroid receptor system in fish compared with in mammals. Here we established a transient transactivation assay using the Japanese medaka, Oryzias latipes, glucocorticoid receptors (olGRs) and mineralocorticoid receptor to analyse their functional properties in a fish. We found that olGR2 was highly responsive to glucocorticoids, similar to the human GR, whereas the olGR1 subtype was minimally responsive. Thus, olGR2 most likely mediates glucocorticoid signaling in medaka. We further tested crosstalk between GRs and other steroid hormones, and found that progestins could activate or inactivate olGR2‐mediating transcription, depending on the presence or absence of cortisol. The transactivation assays developed for medaka GRs provide tools to gain useful insights into corticosteroid signaling in fish and for in vitro screening of environmental substances activating GRs. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-23T08:42:52.17801-05:0
      DOI: 10.1002/jat.3020
  • Cytotoxic and apoptotic activities of the plancitoxin I from the venom of
           crown‐of‐thorns starfish (Acanthaster planci) on A375.S2 cells
    • Authors: Chi‐Chiu Lee; Hernyi Justin Hsieh, Deng‐Fwu Hwang
      Abstract: This study reports on a cytotoxic toxin derived from the venom of the crown‐of‐thorns starfish Acanthaster planci (CAV). The protein toxin was isolated through both ion‐exchange and gel‐filtration chromatography, and characterized by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) and mass spectrum analyzes. The CAV was identified as plancitoxin I protein. The mechanistic role of the CAV toxin was explored in human malignant melanoma A375.S2 cell death. The results indicated that after incubation with CAV toxin, cells significantly decreased in A375.S2 cell viability and increased in the lactate dehydrogenase (LDH) level in a dose‐dependent manner. The assays indicated that CAV toxin promoted reactive oxygen species (ROS) production, induced nitric oxide (NO) formation, lost mitochondrial membrane potential (ΔΨm) and induced inter‐nucleosomal DNA fragmentation in A375.S2 cells. The molecular cytotoxicity of the CAV toxin was tested through evaluation of the apoptosis/necrosis ratio by double staining with annexin V‐FITC and a propidium iodide (PI) assay. The results suggested that CAV toxin induced a cytotoxic effect in A375.S2 cells via the apoptotic procedure, and may be associated with the regulation of the p38 pathways. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-21T09:24:19.994647-05:
      DOI: 10.1002/jat.3034
  • Elevated levels of antibodies against xenobiotics in a subgroup of healthy
    • Authors: Aristo Vojdani; Datis Kharrazian, Partha Sarathi Mukherjee
      Abstract: In spite of numerous research efforts, the exact etiology of autoimmune diseases remains largely unknown. Genetics and environmental factors, including xenobiotics, are believed to be involved in the induction of autoimmune disease. Some environmental chemicals, acting as haptens, can bind to a high‐molecular‐weight carrier protein such as human serum albumin (HSA), causing the immune system to misidentify self‐tissue as an invader and launch an immune response against it, leading to autoimmunity. This study aimed to examine the percentage of blood samples from healthy donors in which chemical agents mounted immune challenges and produced antibodies against HSA‐bound chemicals. The levels of specific antibodies against 12 different chemicals bound to HSA were measured by ELISA in serum from 400 blood donors. We found that 10% (IgG) and 17% (IgM) of tested individuals showed significant antibody elevation against aflatoxin‐HSA adduct. The percentage of elevation against the other 11 chemicals ranged from 8% to 22% (IgG) and 13% to 18% (IgM). Performance of serial dilution and inhibition of the chemical–antibody reaction by specific antigens but not by non‐specific antigens were indicative of the specificity of these antibodies. Although we lack information about chemical exposure in the tested individuals, detection of antibodies against various protein adducts may indicate chronic exposure to these chemical haptens in about 20% of the tested individuals. Currently the pathological significance of these antibodies in human blood is still unclear, and this protein adduct formation could be one of the mechanisms by which environmental chemicals induce autoimmune reactivity in a significant percentage of the population. Copyright © 2014. The
      Authors . Journal of Applied Toxicology Published by John Wiley & Sons Ltd.
      PubDate: 2014-07-18T04:30:19.876942-05:
      DOI: 10.1002/jat.3031
  • Reactive oxygen species‐dependent JNK downregulated
           olaquindox‐induced autophagy in HepG2 cells
    • Authors: Dongxu Zhao; Congcong Wang, Shusheng Tang, Chaoming Zhang, Shen Zhang, Yan Zhou, Xilong Xiao
      Abstract: Autophagy plays an important role in response to intracellular and extracellular stress to sustain cell survival. However, dysregulated or excessive autophagy may lead to cell death, known as “type II programmed cell death,” and it is closely associated with apoptosis. In our previous study, we proposed that olaquindox induced apoptosis of HepG2 cells through a caspase‐9 dependent mitochondrial pathway. In this study, we investigated autophagy induced by olaquindox and explored the crosstalk between apoptosis and autophagy in olaquindox‐treated HepG2 cells. Olaquindox‐induced autophagy was demonstrated by the accumulation of monodansylcadervarine, as well as elevated expression of autophagy‐related MAP‐LC3 and Beclin 1 proteins. The autophagy inhibitor 3‐methyladenine significantly increased the apoptotic rate induced by olaquindox, which was correlated with increased ratio of Bax/Bcl‐2. The further studies showed that olaquindox increased the levels of reactive oxygen species (ROS), and antioxidant N‐acetyl‐L‐cysteine (NAC) effectively blocked the accumulation of ROS but failed to block autophagy. Moreover, olaquindox induced the activation of c‐Jun N‐terminal protein kinase (JNK), and JNK inhibitor SP600125 failed to block autophagy. Instead, olaquindox‐induced autophagy was enhanced by NAC or SP600125. Meanwhile, JNK activation was remarkably blocked by NAC, indicating that ROS may be the upstream signaling molecules of JNK activation and involved in the negative regulation of olaquindox‐induced autophagy. These results suggest that olaquindox induces autophagy in HepG2 cells and that olaquindox‐induced apoptosis can be enhanced by 3‐methyladenine. Olaquindox‐induced autophagy in HepG2 cells is upregulated by Beclin 1 but downregulated by ROS‐dependent JNK. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-18T04:30:06.618358-05:
      DOI: 10.1002/jat.3022
  • Inhibitory effect of apocynin on methylglyoxal‐mediated glycation in
           osteoblastic MC3T3‐E1 cells
    • Authors: Kwang Sik Suh; Sang Youl Rhee, Young Seol Kim, Eun Mi. Choi
      Abstract: Methylglyoxal (MG), a highly reactive metabolite of hyperglycemia, can enhance protein glycation, oxidative stress or inflammation. The present study investigated the effects of apocynin on the mechanisms associated with MG toxicity in osteoblastic MC3T3‐E1 cells. Pretreatment of MC3T3‐E1 cells with apocynin prevented the MG‐induced protein glycation and formation of intracellular reactive oxygen species and mitochondrial superoxide in MC3T3‐E1 cells. In addition, apocynin increased glutathione levels and restored the activity of glyoxalase I inhibited by MG. These findings suggest that apocynin provide a protective action against MG‐induced cell damage by reducing oxidative stress and by increasing the MG detoxification system. Apocynin treatment decreased the levels of proinflammatory cytokines such as tumor necrosis factor‐α and interleukin‐6 induced by MG. Additionally, the nitric oxide level reduced by MG was significantly increased by apocynin. These findings indicate that apocynin might exert its therapeutic effects via upregulation of glyoxalase system and antioxidant activity. Taken together, apocynin may prove to be an effective treatment for diabetic osteopathy. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-18T04:30:03.532821-05:
      DOI: 10.1002/jat.3016
  • Renal cells exposed to cadmium in vitro and in vivo: normalizing gene
           expression data
    • Authors: A. R. Nair; K. Smeets, E. Keunen, W.‐K. Lee, F. Thévenod, E. Van Kerkhove, A. Cuypers
      Abstract: Cadmium (Cd) is a toxic metal with a long half‐life in biological systems. This half‐life is partly as a result of metallothioneins (MTs), metal‐binding proteins with a high affinity for Cd. The high retention properties of the kidneys reside in proximal tubular cells that possess transport mechanisms for Cd‐MT uptake, ultimately leading to more Cd accumulation. Researchers have studied MT–metal interactions using various techniques including quantitative real‐time PCR (qPCR), an efficient tool for quantifying gene expression. Often a poor choice of reference genes, which is represented by their instability and condition dependency, leads to inefficient normalization of gene expression data and misinterpretations. This study demonstrates the importance of an efficient normalization strategy in toxicological research. A selection of stable reference genes was proposed in order to acquire reliable and reproducible gene quantification under metal stress using MT expression as an example. Moreover, in vitro and in vivo setups were compared to identify the influence of toxicological compounds in function of the experimental design. This study shows that glyceraldehyde‐3‐phosphate dehydrogenase (Gapdh), tyrosine monooxygenase/tryptophan5‐monooxygenase activation‐protein, zeta polypeptide (Ywhaz) and beta‐actin (Actb) are the most stable reference genes in a kidney proximal tubular cell line exposed to moderate and high Cd concentrations, applied as CdCl2. A slightly different sequence in reference gene stability was found in renal cells isolated from rats in vivo exposed to Cd. It was further shown that three reference genes are required for efficient normalization in this experimental setup. This study demonstrates the importance of an efficient normalization strategy in toxicological research. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-18T04:29:56.735907-05:
      DOI: 10.1002/jat.3047
  • Toxicity of new emerging pollutant tris‐(2,3‐dibromopropyl)
           isocyanurate on BALB/c mice
    • Authors: Juan Li; Xu Zhang, Jieqing Bao, Yuchen Liu, Junfeng Li, Jia Li, Yong Liang, Jie Zhang, Aiqian Zhang
      Abstract: The emerging heterocyclic brominated flame retardant tris‐(2,3‐dibromopropyl) isocyanurate (TBC), widely used in reinforced plastics, has demonstrated toxicity to fish. However, little is known about its toxicity in rodents. This study aims to determine the effect of TBC on growth, biochemical parameters in serum, organs and related gene expression of both male and female BALB/c mice after gastro‐gavage administration of 0, 2, 10 and 50 mg kg−1 TBC for 28 days. Results indicated that exposure to TBC had no effects on basic growth and food intake of mice, but significantly increased serum alanine aminotransferase levels in male mice. Histopathological analyses showed that focal necrosis (2, 10 and 50 mg kg−1 TBC‐exposed groups) and ballooning degeneration (10 and 50 mg kg−1 TBC‐exposed groups) were found in mouse liver, whereas transmission electron microscopy revealed dose‐dependent hepatocyte apoptosis, mitochondrial degeneration and endoplasmic reticulum dilation. Histopathological and ultrastructural assessments in the lung showed dose‐dependent hyperplasia of pulmonary alveolar epithelium, bronchial congestion, infiltration of inflammatory cells and mitochondrial swelling following TBC exposure. Our results also indicated that mitochondria are one of the major target cytoplasmic organelles for TBC, suggesting that damage in mitochondria is one of the pathways that led to toxic effects in the liver and lung of TBC‐treated groups. Moreover, TBC effectively activated the gene expression of p53 in mice liver. Our findings provide strong evidence that TBC induces significant toxicity in mice organs, especially in liver and lung, which play vital roles in detoxification and gas exchange, respectively. This research will contribute to characterize the toxic effects of TBC, which was introduced as one of the candidates for brominated flame retardant replacement. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-17T23:01:48.126209-05:
      DOI: 10.1002/jat.3026
  • Toxicity profiles and solvent–toxicant interference in the planarian
           Schmidtea mediterranea after dimethylsulfoxide (DMSO) exposure
    • Authors: An‐Sofie Stevens; Nicky Pirotte, Michelle Plusquin, Maxime Willems, Thomas Neyens, Tom Artois, Karen Smeets
      Abstract: To investigate hydrophobic test compounds in toxicological studies, solvents like dimethylsulfoxide (DMSO) are inevitable. However, using these solvents, the interpretation of test compound‐induced responses can be biased. DMSO concentration guidelines are available, but are mostly based on acute exposures involving one specific toxicity endpoint. Hence, to avoid solvent–toxicant interference, we use multiple chronic test endpoints for additional interpretation of DMSO concentrations and propose a statistical model to assess possible synergistic, antagonistic or additive effects of test compounds and their solvents. In this study, the effects of both short‐ (1 day) and long‐term (2 weeks) exposures to low DMSO concentrations (up to 1000 µl l−1) were studied in the planarian Schmidtea mediterranea. We measured different biological levels in both fully developed and developing animals. In a long‐term exposure set‐up, a concentration of 500 µl l−1 DMSO interfered with processes on different biological levels, e.g. behaviour, stem cell proliferation and gene expression profiles. After short exposure times, 500 µl l−1 DMSO only affected motility, whereas the most significant changes on different parameters were observed at a concentration of 1000 µl l−1 DMSO. As small sensitivity differences exist between biological levels and developmental stages, we advise the use of this solvent in concentrations below 500 µl l−1 in this organism. In the second part of our study, we propose a statistical approach to account for solvent–toxicant interactions and discuss full‐scale solvent toxicity studies. In conclusion, we reassessed DMSO concentration limits for different experimental endpoints in the planarian S. mediterranea. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-06-25T22:54:19.05537-05:0
      DOI: 10.1002/jat.3011
  • Comparative genotoxicity of nanosilver in human liver HepG2 and colon
           Caco2 cells evaluated by fluorescent microscopy of cytochalasin
           B‐blocked micronucleus formation
    • Authors: Saura C. Sahu; Shambhu Roy, Jiwen Zheng, Jeffrey J. Yourick, Robert L. Sprando
      Abstract: As a consequence of the increased use of silver nanoparticles in food, food contact materials, dietary supplements and cosmetics to prevent fungal and bacterial growth, there is a need for validated rapid screening methods to assess the safety of nanoparticle exposure. This study evaluated two widely used in vitro cell culture models, human liver HepG2 cells and human colon Caco2 cells, as tools for assessing the potential genotoxicity of 20‐nm nanosilver. The average silver nanoparticle size as determined by transmission electron microscopy (TEM) was 20.4 nm. Dynamic light scattering (DLS) analysis showed no large agglomeration of the silver nanoparticles. The silver concentration in a 20‐nm nanosilver solution determined by the inductively coupled plasma–mass spectrometry (ICP‐MS) analysis was 0.962 mg ml−1. Analysis by ICP‐MS and TEM demonstrated the uptake of 20‐nm silver by both HepG2 and Caco2 cells. Genotoxicity was determined by the cytochalasin B‐blocked micronucleus assay with acridine orange staining and fluorescence microscopy. Concentration‐ and time‐dependent increases in the frequency of binucleated cells with micronuclei induced by the nanosilver was observed in the concentration range of 0.5 to 15 µg ml−1 in both HepG2 and Caco2 cells compared with the control. Our results indicated that HepG2 cells were more sensitive than Caco2 cells in terms of micronuclei formation induced by nanosilver exposure. In summary, the results of this study indicate that the widely used in vitro models, HepG2 and Caco2 cells in culture, represent potential screening models for prediction of genotoxicity of silver nanoparticles by in vitro micronucleus assay. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.
      PubDate: 2014-06-06T09:15:06.224268-05:
      DOI: 10.1002/jat.3028
  • Time‐dependent bioaccumulation of distinct rod‐type TiO2
           nanoparticles: Comparison by crystalline phase
    • Authors: Eun-Jung Park; Gwang-Hee Lee, Cheolho Yoon, Min-Sung Kang, Soo Nam Kim, Myung-Haing Cho, Jae-Ho Kim, Dong-Wan Kim
      Abstract: A complete understanding of the interaction between nanoparticles and biological systems, including nanoparticle uptake and distribution and the biological responses, could guide the design of safer and more effective nanoparticles than those currently available. In this study, we compared the distribution in mice over time of two rod‐type titanium dioxide nanoparticles (TiNPs) that feature distinct phases, anatase (ATO) and brookite (BTO). Surface areas of BTO and ATO were estimated to be 102 and 268 m2 g–1, respectively, and negative charge on the surface of ATO was higher than that of BTO in deionized water. Both TiNPs were rapidly distributed into tissues after injection. At 4 weeks after injection, both TiNPs were maximally accumulated in the spleen, followed by the liver, but the total accumulation of ATO in tissues measured in this study was more than that of BTO. Moreover, the cellular antioxidant function was similar although the levels of Ti measured in tissues were distinct between the two TiNPs. Based on these results, we suggest that the fate of TiNPs in the body may differ according to the size and surface charge of the TiNPs even when their shape is the same. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-29T08:38:10.446352-05:
      DOI: 10.1002/jat.3006
  • Vascular endothelial growth factor mRNA levels as a biomarker for
           short‐term N‐butyl‐N‐(4‐hydroxybutyl)
           nitrosamine‐induced rat bladder carcinogenesis bioassay
    • Authors: Shin Wakui; Tomoko Mutou, Hiroyuki Takahashi, Masahiro Ikegami, Hideki Wanibuchi, Shoji Fukushima
      Abstract: Generically, carcinogenic effects of chemicals in bladder carcinogenesis are judged by induction of papillary or nodular (PN) hyperplasia in rats given N‐butyl‐N‐(4‐hydroxybutyl) nitrosamine (BBN) for 4 weeks and the test chemical for 22–28 weeks. However, upregulation of vascular endothelial growth factor (VEGF) begins early in rat BBN bladder carcinogenesis. To establish a short‐term rat bladder carcinogenic bioassay, we analyzed the correlations between VEGF, VEGF mRNA and bladder lesions inductions at 10 and 26 weeks after BBN treatment. Six‐week‐old male Wistar (slc) rats were given 0.05% BBN for 4, 10 or 26 weeks. To avoid individual rat bias, the bladders were investigated by partial cystectomy at 10 weeks and total cystectomy at 26 weeks. After induction, PN hyperplasia and carcinoma in rats increased with the length of BBN treatment and immunohistochemical VEGF expression also increased following carcinogenesis, but the immunoreactivity of individual lesions was quite variable. Moreover, induction of PN hyperplasia at 10 weeks’ BBN treatment was not significantly correlated with that at 26 weeks' treatment; thus, it was not possible to predict the carcinogenic effect due to the induction of PN hyperplasia at 26 weeks' BBN treatment by that at 10 weeks' treatment. However, VEGF mRNA levels of rat bladders at 10 weeks' BBN treatment revealed a strong significant correlation with the incidence of bladder lesions at 26 weeks' treatment. Here, we suggest that quantitative VEGF mRNA levels are a good biomarker for a short‐term BBN‐induced bioassay for rat bladder carcinogenesis. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-28T09:25:42.146279-05:
      DOI: 10.1002/jat.3021
  • Developmental toxicity assay using high content screening of zebrafish
    • Authors: Susan Lantz‐McPeak; Xiaoqing Guo, Elvis Cuevas, Melanie Dumas, Glenn D. Newport, Syed F. Ali, Merle G. Paule, Jyotshna Kanungo
      Abstract: Typically, time‐consuming standard toxicological assays using the zebrafish (Danio rerio) embryo model evaluate mortality and teratogenicity after exposure during the first 2 days post‐fertilization. Here we describe an automated image‐based high content screening (HCS) assay to identify the teratogenic/embryotoxic potential of compounds in zebrafish embryos in vivo. Automated image acquisition was performed using a high content microscope system. Further automated analysis of embryo length, as a statistically quantifiable endpoint of toxicity, was performed on images post‐acquisition. The biological effects of ethanol, nicotine, ketamine, caffeine, dimethyl sulfoxide and temperature on zebrafish embryos were assessed. This automated developmental toxicity assay, based on a growth‐retardation endpoint should be suitable for evaluating the effects of potential teratogens and developmental toxicants in a high throughput manner. This approach can significantly expedite the screening of potential teratogens and developmental toxicants, thereby improving the current risk assessment process by decreasing analysis time and required resources. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.
      PubDate: 2014-05-28T09:20:58.316539-05:
      DOI: 10.1002/jat.3029
  • Cardiotoxicity evaluation of anthracyclines in zebrafish (Danio rerio)
    • Authors: Ying Han; Jing‐pu Zhang, Jian‐qin Qian, Chang‐qin Hu
      Abstract: Drug‐induced cardiotoxicity is a leading factor for drug withdrawals, and limits drug efficacy and clinical use. Therefore, new alternative animal models and methods for drug safety evaluation have been given great attention. Anthracyclines (ANTs) are widely prescribed anticancer agents that have a cumulative dose relationship with cardiotoxicity. We performed experiments to study the toxicity of ANTs in early developing zebrafish embryos, especially their effects on the heart. LC50 values for daunorubicin, pirarubicin, doxorubicin (DOX), epirubicin and DOX‐liposome at 72 h post‐fertilization were 122.7 μM, 111.9 μM, 31.2 μM, 108.3 μM and 55.8 μM, respectively. At the same time, zebrafish embryos were exposed to ANTs in three exposure stages and induced incomplete looping of the heart tube, pericardia edema and bradycardia in a dose‐dependent manner, eventually leading to death. DOX caused the greatest heart defects in the treatment stages and its liposome reduced the effects on the heart, while daunorubicin produced the least toxicity. Genes and proteins related to heart development were also identified to be sensitive to ANT exposure and downregulated by ANTs. It revealed ANTs could disturb the heart formation and development. ANTs induced cardiotoxicity in zebrafish has similar effects in mammalian models, indicating that zebrafish may have a potential value for assessment of drug‐induced developmental cardiotoxicity. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-22T16:22:52.444139-05:
      DOI: 10.1002/jat.3007
  • A trivalent approach for determining in vitro toxicology: Examination of
           oxime K027
    • Authors: Adriana Prado; Georg A. Petroianu, Dietrich E. Lorke, Jeremy W. Chambers
      Abstract: Unforeseen toxic effects contribute to compound attrition during preclinical evaluation and clinical trials. Consequently, there is a need to correlate in vitro toxicity to in vivo and clinical outcomes quickly and effectively. We propose an expedited evaluation of physiological parameters in vitro that will improve the ability to predict in vivo toxicity of potential therapeutics. By monitoring metabolism, mitochondrial physiology and cell viability, our approach provides insight to the extent of drug toxicity in vitro. To implement our approach, we used human hepatocellular carcinoma cells (HepG2) and neuroblastoma cells (SH‐SY5Y) to monitor hepato‐ and neurotoxicity of the experimental oxime K027. We utilized a trivalent approach to measure metabolism, mitochondrial stress and induction of apoptosis in 96‐well formats. Any change in these three areas may suggest drug‐induced toxicity in vivo. K027 and pralidoxime, an oxime currently in clinical use, had no effect on glycolysis or oxygen consumption in HepG2 and SH‐SY5Y cells. Similarly, these oximes did not induce oxidant generation nor alter mitochondrial membrane potential. Further, K027 and pralidoxime failed to activate effector caspases, and these oximes did not alter viability. The chemotherapeutic agent, docetaxel, negatively affected metabolism, mitochondrial physiology and viability. Our studies present a streamlined high‐throughput trivalent approach for predicting toxicity in vitro, and this approach reveals that K027 has no measurable hepatotoxicity or neurotoxicity in vitro, which correlates with their in vivo data. This approach could eliminate toxic drugs from consideration for in vivo preclinical evaluation faster than existing toxicity prediction panels and ultimately prevent unnecessary experimentation. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-22T16:22:49.472421-05:
      DOI: 10.1002/jat.3013
  • Synergistic cytotoxicity and DNA strand breaks in cells and plasmid DNA
           exposed to uranyl acetate and ultraviolet radiation
    • Authors: Janice Wilson; Mary C. Zuniga, Filbert Yazzie, Diane M. Stearns
      Abstract: Depleted uranium (DU) has a chemical toxicity that is independent of its radioactivity. The purpose of this study was to explore the photoactivation of uranyl ion by ultraviolet (UV) radiation as a chemical mechanism of uranium genotoxicity. The ability of UVB (302 nm) and UVA (368 nm) radiation to photoactivate uranyl ion to produce single strand breaks was measured in pBR322 plasmid DNA, and the presence of adducts and apurinic/apyrimidinic sites that could be converted to single strand breaks by heat and piperidine was analyzed. Results showed that DNA lesions in plasmid DNA exposed to UVB‐ or UVA‐activated DU were only slightly heat reactive, but were piperidine sensitive. The cytotoxicity of UVB‐activated uranyl ion was measured in repair‐proficient and repair‐deficient Chinese hamster ovary cells and human keratinocyte HaCaT cells. The cytotoxicity of co‐exposures of uranyl ion and UVB radiation was dependent on the order of exposure and was greater than co‐exposures of arsenite and UVB radiation. Uranyl ion and UVB radiation were synergistically cytotoxic in cells, and cells exposed to photoactivated DU required different DNA repair pathways than cells exposed to non‐photoactivated DU. This study contributes to our understanding of the DNA lesions formed by DU, as well as their repair. Results suggest that excitation of uranyl ion by UV radiation can provide a pathway for uranyl ion to be chemically genotoxic in populations with dermal exposures to uranium and UV radiation, which would make skin an overlooked target organ for uranium exposures. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-15T09:03:01.061665-05:
      DOI: 10.1002/jat.3015
  • Endocrine disruption effects of p,p′‐DDE on juvenile zebrafish
    • Authors: Marta Sofia Monteiro; Maria Pavlaki, Augusto Faustino, Alexandra Rêma, Mariana Franchi, Letícia Gediel, Susana Loureiro, Inês Domingues, Jaime Rendón von Osten, Amadeu Mortágua Velho Maia Soares
      Abstract: The persistent organic pollutant p,p′‐DDE, the major metabolite of the insecticide DDT, has displayed evidence of endocrine disruption through the inhibition of androgen binding to androgen receptors in different species. Although p,p′‐DDE was continuously detected in wild fish with abnormal gonad development such as intersex, little is known about its mode of action during gonad development in fish. To elucidate the potential endocrine effects of this pollutant in zebrafish (Danio rerio), juveniles (30 days post hatch) were exposed to p,p′‐DDE during the critical window of sexual differentiation. Fish were exposed to sublethal concentrations ranging from 0.01 to 20 µg l–1 over 14 days and were maintained in control water for an additional 4 months. As core endpoints, the vitellogenin (vtg) concentration was measured at the end of exposure, and sex ratio and the gonadosomatic index were assessed 4 months after the end of exposure. An increase in vtg production in whole body homogenate was observed in fish exposed to 0.2 and 2.0 µg l–1 p,p′‐DDE. No significant differences were displayed in morphological parameters such as the gonadosomatic index of males and females or sex ratio. However, exposed females presented histopathological changes that include the reduction of the number of mature oocytes, which might impair their successful reproduction. These results demonstrate the ability of p,p′‐DDE to cause endocrine disruption in zebrafish exposed during gonad differentiation of juvenile specimens. Furthermore, vtg induction by p,p′‐DDE in juvenile zebrafish arises as a predictive marker for adverse effects of this DDT metabolite on the ovarian function of female zebrafish. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-15T08:55:11.837345-05:
      DOI: 10.1002/jat.3014
  • Dimethylarsinic acid (DMAv) changed the expressions of proliferative
           related factors and secretion of inflammatory cytokines in rat bladder
    • Authors: Zhang Lin; Liu Shengnan, Wang Fei, Song Yingli, Sun Qingshan, Sheng Wei, Xi. Shuhua, Sun Guifan
      Abstract: Dimethylarsinic acid (DMAV), the major urinary metabolite of inorganic arsenic, is a urinary bladder carcinogen and bladder tumor promoter in adult rats. Increased urothelial cellular proliferation has been considered as an earlier phenotype in DMAV‐induced bladder carcinogenesis. The present study examined the ultrastructural changes of bladder epithelial cells and expressions of proliferation factors, as well as the secretion of inflammatory cytokines in rats exposed to DMAV for 10 weeks by transmission electron microscopy (TEM), qRT‐PCR, immunohistochemical staining and ELISA methods. The results showed that DMAV administered in the drinking water produced urothelial cytotoxicity and ultrastructural changes in rats. PCNA, cyclin D1 and COX‐2 mRNA expressions and immunoreactivities were elevated in bladder urothelium. In addition, 200 ppm DMAV treatment increased the transforming growth factor‐beta 1 (TGF‐β1) secretion and decreased tumor necrosis factor‐alpha (TNF)‐α level in the urine of rats. These data suggest that chronic inflammation, bladder epithelium lesions and proliferation might be the basic process of the chronic toxicity effects in DMAV‐treated rats. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-05-15T08:46:33.106421-05:
      DOI: 10.1002/jat.3001
  • Usefulness of urinary kidney injury molecule‐1 (Kim‐1) as a
           biomarker for cisplatin‐induced sub‐chronic kidney injury
    • Authors: Ken‐ichiro Nan‐ya; Masatomo Kajihara, Natsuki Kojima, Masakuni Degawa
      Abstract: We explored biomarkers suitable for monitoring sub‐chronic kidney injury using the three rat models of cisplatin (CDDP)‐induced kidney injury, which were designed to extend the current knowledge beyond the sub‐acute exposure period. In the pilot study, a single intravenous administration of 1.5 mg kg–1 CDDP to rats was confirmed to result in no histopathological changes. Subsequently, CDDP was intravenously administered to rats at a dose of 1.5 mg kg–1 for 4 days at 24‐h intervals (Experimental model 1) and for up to 10 weeks at weekly intervals (Experimental models 2 and 3), and the changes in blood and urine components, such as recently recommended urinary biomarkers (Kim‐1, clusterin and so on) and traditional blood biomarkers (blood urea nitrogen and serum creatinine), were examined together with the histopathological changes in renal tissues during the development of the kidney injury in each model. In these experimental models, a significant increase in urinary Kim‐1 was observed prior to the histopathological changes in renal tissues, and these changes were retained after the adverse histopathological changes. Significant changes in all of the other urinary biomarkers examined occurred along with the histopathological changes. In addition, the increase in urinary Kim‐1 after weekly treatment with CDDP for 4 weeks was reduced in a time‐dependent manner after cessation of the drug. The present findings indicate that urinary Kim‐1 is the most useful biomarker for CDDP‐induced rat sub‐chronic kidney injury among the biomarkers examined. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-04-16T05:43:48.985818-05:
      DOI: 10.1002/jat.2999
  • Evaluation of developmental toxicity using undifferentiated human
           embryonic stem cells
    • Authors: Eui‐Man Jung; Yeo‐ul Choi, Hong‐Seok Kang, Hyun Yang, Eui‐Ju Hong, Beum‐Soo An, Jun‐young Yang, Ki. Hwan Choi, Eui‐Bae Jeung
      Abstract: An embryonic stem cell test (EST) has been developed to evaluate the embryotoxic potential of chemicals with an in vitro system. In the present study, novel methods to screen toxic chemicals during the developmental process were evaluated using undifferentiated human embryonic stem (hES) cells. By using surface marker antigens (SSEA‐4, TRA‐1‐60 and TRA‐1‐81), we confirmed undifferentiated conditions of the used hES cells by immunocytochemistry. We assessed the developmental toxicity of embryotoxic chemicals, 5‐fluorouracil, indomethacin and non‐embryotoxic penicillin G in different concentrations for up to 7 days. While expressions of the surface markers were not significantly affected, the embryotoxic chemicals influenced their response to pluripotent ES cell markers, such as OCT‐4, NANOG, endothelin receptor type B (EDNRB), secreted frizzled related protein 2 (SFRP2), teratocarcinoma‐derived growth factor 1 (TDGF1), and phosphatase and tensin homolog (PTEN). Most of the pluripotent ES cell markers were down‐regulated in a dose‐dependent manner after treatment with embryotoxic chemicals. After treatment with 5‐fluorouracil, indomethacin and penicillin G, we observed a remarkable convergence in the degree of up‐regulation of development, cell cycle and apoptosis‐related genes by gene expression profiles using an Affymetrix GeneChips. Taken together, these results suggest that embryotoxic chemicals have cytotoxic effects, and modulate the expression of ES cell markers as well as development‐, cell cycle‐ and apoptosis‐related genes that have pivotal roles in undifferentiated hES cells. Therefore, we suggest that hES cells may be useful for testing the toxic effects of chemicals that could impact the embryonic developmental stage. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-04-16T05:43:46.307596-05:
      DOI: 10.1002/jat.3010
  • Serum enhanced cytokine responses of macrophages to silica and iron oxide
           particles and nanomaterials: a comparison of serum to lung lining fluid
           and albumin dispersions
    • Authors: David M. Brown; Helinor Johnston, Eva Gubbins, Vicki Stone
      Abstract: The potential hazard to humans exposed to nanomaterials such as silica and iron oxide was investigated using an in vitro macrophage cell culture system. Amorphous silica and iron oxide particles and nanomaterials (NMs) were dispersed in cell culture medium supplemented with either bovine serum albumin (BSA), lung lining fluid (LLF) or serum, in order to mimic the body fluids encountered during different routes of exposure in the body. End points investigated included macrophage viability and cytokine production. Silica NMs and particles (50 and 200 nm, respectively) were unmodified (plain) or aminated (NH2). Iron oxide NMs and particles, Fe3O4 45 nm and Fe2O3 280 nm were also used in this study. Silica particles and NMs induced a dose‐dependent increase in cytotoxicity as measured by lactate dehydrogenase (LDH) release. Serum enhanced silica‐induced interleukin (IL)‐6, IL‐10, IL‐1β and MCP‐1 release, whereas albumin partially inhibited MCP‐1 release. Aminated silica, 50 nm was more potent than the 200‐nm particles at inducing monocyte chemoattractant protein‐1 (MCP‐1) production when dispersed in medium or LLF, suggesting a size specific effect for these particles and this cytokine. Iron oxide particles were relatively inert compared with the silica particles and NMs; however, serum and albumin did affect cytokine release in some treatments. In conclusion, the data suggests that serum, compared with medium, BSA and LLF is very potent at enhancing macrophage responses to silica and iron oxide particles and NMs. Size was only influential in LLF for a limited number of parameters, whereas surface chemistry was not of consequence in this in vitro macrophage system. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-04-16T05:29:44.649109-05:
      DOI: 10.1002/jat.2998
  • In vitro exposure of DE‐71, a penta‐PBDE mixture, on immune
           endpoints in bottlenose dolphins (Tursiops truncatus) and B6C3F1 mice
    • Authors: Jena R. Wirth; Margie M. Peden‐Adams, Natasha D. White, Gregory D. Bossart, Patricia A. Fair
      Abstract: Polybrominated diphenyl ethers (PBDEs) are an emerging contaminant of concern with low level exposures demonstrating toxicity in laboratory animals and wildlife, although immunotoxicity studies have been limited. Bottlenose dolphin peripheral blood leukocytes (PBLs) and mouse splenocytes were exposed to environmentally relevant DE‐71 (a penta‐PBDE mixture) concentrations (0–50 µg ml−1) in vitro. Natural killer (NK) cell activity and lymphocyte (B and T cell) proliferation were evaluated using the parallelogram approach for risk assessment. This study aimed to substantiate results from field studies with dolphins, assess the sensitivities between the mouse model and dolphins, and to evaluate risk using the parallelogram approach. In mouse cells, NK cell activity increased at in vitro doses 0.05, 0.5 and 25 µg DE‐71 ml−1, whereas proliferation was not modulated. In dolphin cells, NK cell activity and lymphocyte proliferation was not altered after in vitro exposure. In vitro exposure of dolphin PBLs to DE‐71 showed similar results to correlative field studies; NK cell activity in mice was more sensitive to in vitro exposure than dolphins, and the parallelogram approach showed correlation with all three endpoints to predict risk in bottlenose dolphins. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-04-07T02:26:12.183666-05:
      DOI: 10.1002/jat.3008
  • Impact of acute arsenic and cadmium exposure on the expression of two
           haeme oxygenase genes and other antioxidant markers in common carp
           (Cyprinus carpio)
    • Authors: Zsanett Jancsó; Edit Hermesz
      Abstract: The aim was to study the effects of cadmium (Cd) and arsenic (As) on haeme oxygenases (HOs) and other oxidative stress biomarkers, and their roles in macromolecule damage in liver and kidney of common carp (Cyprinus carpio L.). HOs play a critical role in the defence system against oxidative damage, producing biliverdin and carbon monoxide with important free radical scavenging properties. However, increased HO activity in haeme degradation may also lead to a pro‐oxidant effect through the liberation of Fe‐modifying Cd and As toxicity. The response of an organism to exposure to toxic metals is in many cases brought about by changes at the level of gene expression. In this study, the genes ho‐1 and ho‐2 of the common carp were identified, and the changes in gene expressions were analysed from the aspect of Cd and As accumulation. Both ho‐1 and ho‐2 are transcriptionally induced by Cd and As, but their inductions differ in time course, dose response and tissue specificity. The expression of ho1 was mostly affected by As, primarily in the liver (45‐fold), whereas it was enhanced with higher efficacy by Cd in the kidney (25‐fold). The cellular redox status and the damage of lipid molecules were monitored via the ratio of reduced to oxidized glutathione, the levels of H2O2 and lipid peroxidation, and the activities of superoxide dismutase (SOD) and catalase (CAT). Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-04-07T02:26:09.502028-05:
      DOI: 10.1002/jat.3000
  • Usefulness of optical coherence tomography to detect central serous
           chorioretinopathy in monkeys
    • Authors: Hyun‐Kyu Park; Woori Jo, Hyun‐Ji Choi, Bongcheol Kim, Gilnam Lee, Jeongbeob Seo, Suk Young Cho, Choung‐Soo Kim, Eun Kyung Choi, Jung Jin Hwang, Joo Yong Lee, Young Hee Yoon, Woo‐Chan Son
      Abstract: Many systemic drugs can induce ocular toxicity and several ocular side‐effects have been identified in clinical studies. However, it is difficult to detect ocular toxicity in preclinical studies because of the lack of appropriate evaluation methods. Optical coherence tomography (OCT) is useful because it can provide real‐time images throughout a study period, whereas histopathology only provides images of sacrificed animals. Using OCT alongside histopathology, attempts were made to find effective approaches for screening of drug‐induced ocular toxicity in monkeys. Such approaches could be used in preclinical studies prior to human trials. Six male cynomolgus monkeys (Macaca fascicularis Raffles) were orally administered one of six candidate MAPK/ERK kinase (MEK) inhibitors. Central serous chorioretinopathy, a known side‐effect of such inhibitors, was identified in four monkeys by OCT. Artifacts generated during tissue processing meant that histopathology could not detect edematous changes. Thus, OCT is a useful tool to detect ocular toxicity which cannot be detected by histopathology in preclinical studies. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-03-28T03:47:29.247172-05:
      DOI: 10.1002/jat.3009
  • Involvement of immune‐ and inflammatory‐related factors in
           flucloxacillin‐induced liver injury in mice
    • Authors: Shohei Takai; Satonori Higuchi, Azusa Yano, Koichi Tsuneyama, Tatsuki Fukami, Miki Nakajima, Tsuyoshi Yokoi
      Abstract: Drug‐induced liver injury (DILI) is a serious problem in pre‐clinical stages of drug development and clinical pharmacotherapy, but the pathogenesis of DILI has not been elucidated. Flucloxacillin (FLX), which is a β‐lactam antibiotic of the penicillin class that is used widely in Europe and Australia, rarely causes DILI. Clinical features suggest that FLX‐induced liver injury is caused by immune‐ and inflammatory‐related factors, but the mechanism of FLX‐induced liver injury is unknown. The purpose of this study was to elucidate the mechanisms of FLX‐induced liver injury in vivo. Plasma alanine aminotransferase, aspartate aminotransferase and total‐bilirubin levels were significantly elevated in FLX‐administered mice [1000 mg kg–1, intraperitoneally (i.p.)]. Toll‐like receptor 4 (TLR4) ligands, such as high‐mobility group box 1 (HMGB1) and S100A8/A9, were significantly increased in FLX‐administered mice, and inflammatory factors, such as interleukin (IL)‐1β, tumor necrosis factor‐alpha (TNF‐α), macrophage inflammatory protein (MIP)‐2, CXC chemokine‐ligand‐1 (CXCL1) and monocyte chemoattractant protein (MCP)‐1, were also significantly elevated. IL‐17‐related transcriptional factors and cytokines were increased, and the administration of recombinant IL‐17 (2 mg per body weight, i.p.) resulted in an exacerbation of the FLX‐induced liver injury. TLR4‐associated‐signal transduction may be involved in FLX‐induced liver injury, and IL‐17 is an exacerbating factor. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-03-20T06:29:38.953028-05:
      DOI: 10.1002/jat.3002
  • Molecular biomarkers of phospholipidosis in rat blood and heart after
           amiodarone treatment
    • Authors: Nicola Bocchini; Mery Giantin, Federica Crivellente, Serena Ferraresso, Ivo Faustinelli, Mauro Dacasto, Patrizia Cristofori
      Abstract: Phospholipidosis (PLD) is characterized by an intracellular accumulation of phospholipids in lysosomes and concurrent development of concentric lamellar bodies. It is induced in humans and in animals by drugs with a cationic amphiphilic structure. The purpose of the present study was to identify a set of molecular biomarkers of PLD in rat blood and heart, hypothetically applicable in preclinical screens within the drug development process. A toxicological study was set up in rats orally treated up to 11 days with 300 mg kg–1 per day–1 amiodarone (AMD). Light and transmission electron microscopy investigations were performed to confirm the presence of lamellar bodies indicative of phospholipid accumulation. The effects of AMD upon the transcriptome of these tissues were estimated using DNA microarray technology. Microarray data analysis showed that a total of 545 and 8218 genes were modulated by AMD treatment in heart and blood, respectively. Some genes implicated in the phospholipid accumulation in cells, such as phospholipase A2, showed similar alterations of gene expression. After transcriptome criteria of analysis and target selection, including also the involvement in the onset of PLD, 7 genes (Pla2g2a, Pla2g7, Gal, Il1b, Cebpb, Fcgr2b, Acer 2) were selected as candidate biomarkers of PLD in heart and blood tissues, and their potential usefulness as a sensitive screening test was screened and confirmed by quantitative Real‐Time PCR analysis. Collectively, these data underscore the importance of transcriptional profiling in drug discovery and development, and suggest blood as a surrogate tissue for possible phospholipid accumulation in cardiomyocytes. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-18T00:25:23.937611-05:
      DOI: 10.1002/jat.2992
  • Preliminary safety evaluation of a taurocholate‐conjugated
           low‐molecular‐weight heparin derivative (LHT7): a potent
           angiogenesis inhibitor
    • Authors: Farzana Alam; Seung Woo Chung, Seung Rim Hwang, Ji‐young Kim, Jooho Park, Hyun Tae Moon, Youngro Byun
      Abstract: In our previous studies, taurocholic acid (TA)‐conjugated low‐molecular‐weight heparin derivative (LHT7) has been proven to be a potent anti‐angiogenic agent by demonstrated successful blockage capability of vascular endothelial growth factors (VEGF). Preliminary safety evaluations were conducted based on its mechanism of action and chemical behavior. For this purpose, acute toxicity study, and hematological and serological evaluations were carried out. Additionally, in order to evaluate mechanism‐related side effects, both blood pressure and the occurrence of proteinuria were measured using a treatment regime of multiple high doses of LHT7 in a biodistribution study. LD50 values for LHT7 in female and male mice were 56.9 and 64.7 mg kg–1 doses, respectively. There were no vital fluctuations in the serological and hematological parameters, except for the elevated levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) at 100 and 200 mg kg–1 doses of LHT7, representing vital changes in the liver function. Moreover, the results of mechanism‐related studies showed that blood pressure at 50 mg kg–1 did not change but showed elevated levels of protein in urine. In the biodistribution study, a slight accumulation of LHT7 in the kidney and the liver were observed at the 50 mg kg–1 repeated dose owing to the presence of bile acid. No fatal damage was observed in this study; most observations were related to the chemical composition or the mechanism of action of the material. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-16T21:58:07.934802-05:
      DOI: 10.1002/jat.2995
  • Interaction of citrate‐coated silver nanoparticles with earthworm
           coelomic fluid and related cytotoxicity in Eisenia andrei
    • Authors: Jin Il Kwak; Woo‐Mi Lee, Shin Woong Kim, Youn‐Joo An
      Abstract: Understanding the interaction of nanoparticles with biological fluid is important for predicting the behavior and toxicity of nanoparticles in living systems. The earthworm Eisenia andrei was exposed to citrate‐coated silver nanoparticles (cAgNPs), and the interaction of cAgNPs with earthworm coelomic fluid (ECF), the cytotoxicity of cAgNPs in earthworm coelomocytes was assessed. The neutral red retention assay showed a reduction in lysosomal stability after exposure. The toxicity of silver ions dissolved from cAgNPs in the soil medium was not significant. The aggregation and dissolution of cAgNPs increased in ECF, which contains various electrolytes that alter the properties of nanoparticles, and their subsequent toxicity. Microscopic and dissolution studies demonstrated that the aggregation of cAgNPs rapidly increased, and readily dissolved in ECF. The bioavailability of cAgNPs to earthworms induced lysosomal cytotoxicity. This is the first report to test the interaction and lysosomal cytotoxicity of nanoparticles in earthworm biofluids. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-16T21:47:10.518445-05:
      DOI: 10.1002/jat.2993
  • Carbon‐based nanomaterials accelerate arteriolar thrombus formation
           in the murine microcirculation independently of their shape
    • Authors: Martin Holzer; Peter Bihari, Marc Praetner, Bernd Uhl, Christoph Reichel, Janos Fent, Minnamari Vippola, Susan Lakatos, Fritz Krombach
      Abstract: Although carbon‐based nanomaterials (CBNs) have been shown to exert prothrombotic effects in microvessels, it is poorly understood whether CBNs also have the potential to interfere with the process of leukocyte‐endothelial cell interactions and whether the shape of CBNs plays a role in these processes. Thus, the aim of this study was to compare the acute effects of two differently shaped CBNs, fiber‐shaped single‐walled carbon nanotubes (SWCNT) and spherical ultrafine carbon black (CB), on thrombus formation as well as on leukocyte‐endothelial cell interactions and leukocyte transmigration in the murine microcirculation upon systemic administration in vivo. Systemic administration of both SWCNT and CB accelerated arteriolar thrombus formation at a dose of 1 mg kg–1 body weight, whereas SWCNT exerted a prothrombotic effect also at a lower dose (0.1 mg kg–1 body weight). In vitro, both CBNs induced P‐selectin expression on human platelets and formation of platelet‐granulocyte complexes. In contrast, injection of fiber‐shaped SWCNT or of spherical CB did not induce leukocyte–endothelial cell interactions or leukocyte transmigration. In vitro, both CBNs slightly increased the expression of activation markers on human monocytes and granulocytes. These findings suggest that systemic administration of CBNs accelerates arteriolar thrombus formation independently of the CBNs' shape, but does not induce leukocyte–endothelial cell interactions or leukocyte transmigration. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-14T07:58:14.20484-05:0
      DOI: 10.1002/jat.2996
  • Development and utilization of an ex vivo bromodeoxyuridine local lymph
           node assay protocol for assessing potential chemical sensitizers
    • Authors: W. C. Williams; C. Copeland, E. Boykin, S. J. Quell, D. M. Lehmann
      Abstract: The murine local lymph node assay (LLNA) is widely used to identify chemicals that may cause allergic contact dermatitis. Exposure to a dermal sensitizer results in proliferation of local lymph node T cells, which has traditionally been measured by in vivo incorporation of [3H]methyl thymidine. A more recent non‐isotopic variation of the assay utilizes bromodeoxyuridine (BrdU) incorporation in vivo. To further improve the utility of this assay, we developed an ex vivo BrdU labeling procedure eliminating the need for in vivo injections. The results of this assay correctly identified a strong sensitizer (i.e., trimellitic anhydride) as well as weak/moderate sensitizers (i.e., eugenol, cinnamaldehyde and hexylcinnaminic aldehyde). As anticipated, neither non‐sensitizers isopropanol and lactic acid nor the false negative chemical nickel II sulfate hexahydrate induced a positive threshold response in the assay. The results of this assay are in close agreement with those of the in vivo LLNA:BrdU‐enzyme‐linked immunosorbent assay labeling procedure. We also used the ex vivo BrdU LLNA procedure to evaluate ammonium hexachloroplatinate, ammonium tetrachloroplatinate and cis‐diamminedichloroplatinum(II) and the assay correctly identified them as sensitizers based on the calculation of EC2 values. We conclude that this ex vivo BrdU labeling method offers predictive capacity comparable to previously established LLNA protocols while eliminating animal injections and the use of radioisotope. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.
      PubDate: 2014-02-14T07:42:07.38687-05:0
      DOI: 10.1002/jat.2983
  • Comparative cytotoxicity of nanosilver in human liver HepG2 and colon
           Caco2 cells in culture
    • Authors: Saura C. Sahu; Jiwen Zheng, Lesley Graham, Lynn Chen, John Ihrie, Jeffrey J. Yourick, Robert L. Sprando
      Abstract: The use of silver nanoparticles in food, food contact materials, dietary supplements and cosmetics has increased significantly owing to their antibacterial and antifungal properties. As a consequence, the need for validated rapid screening methods to assess their toxicity is necessary to ensure consumer safety. This study evaluated two widely used in vitro cell culture models, human liver HepG2 cells and human colon Caco2 cells, as tools for assessing the potential cytotoxicity of food‐ and cosmetic‐related nanoparticles. The two cell culture models were utilized to compare the potential cytotoxicity of 20‐nm silver. The average size of the silver nanoparticle determined by our transmission electron microscopy (TEM) analysis was 20.4 nm. The dynamic light scattering (DLS) analysis showed no large agglomeration of the silver nanoparticles. The concentration of the 20‐nm silver solution determined by our inductively coupled plasma–mass spectrometry (ICP‐MS) analysis was 0.962 mg ml–1. Our ICP‐MS and TEM analysis demonstrated the uptake of 20‐nm silver by both HepG2 and Caco2 cells. Cytotoxicity, determined by the Alamar Blue reduction assay, was evaluated in the nanosilver concentration range of 0.1 to 20 µg ml–1. Significant concentration‐dependent cytotoxicity of the nanosilver in HepG2 cells was observed in the concentration range of 1 to 20 µg ml–1 and at a higher concentration range of 10 to 20 µg ml–1 in Caco2 cells compared with the vehicle control. A concentration‐dependent decrease in dsDNA content was observed in both cell types exposed to nanosilver but not controls, suggesting an increase in DNA damage. The DNA damage was observed in the concentration range of 1 to 20 µg ml–1. Nanosilver‐exposed HepG2 and Caco2 cells showed no cellular oxidative stress, determined by the dichlorofluorescein assay, compared with the vehicle control in the concentration range used in this study. A concentration‐dependent decrease in mitochondria membrane potential in both nanosilver exposed cell types suggested increased mitochondria injury compared with the vehicle control. The mitochondrial injury in HepG2 cells was significant in the concentration range of 1 to 20 µg ml–1, but in Caco2 cells it was significant at a higher concentration range of 10 to 20 µg ml–1. These results indicated that HepG2 cells were more sensitive to nanosilver exposure than Caco2 cells. It is generally believed that cellular oxidative stress induces cytotoxicity of nanoparticles. However, in this study we did not detect any nanosilver‐induced oxidative stress in either cell type at the concentration range used in this study. Our results suggest that cellular oxidative stress did not play a major role in the observed cytotoxicity of nanosilver in HepG2 and Caco2 cells and that a different mechanism of nanosilver‐induced mitochondrial injury leads to the cytotoxicity. The HepG2 and Caco2 cells used this study appear to be targets for silver nanoparticles. The results of this study suggest that the differences in the mechanisms of toxicity induced by nanosilver may be largely as a consequence of the type of cells used. This differential rather than universal response of different cell types exposed to nanoparticles may play an important role in the mechanism of their toxicity. In summary, the results of this study indicate that the widely used in vitro models, HepG2 and Caco2 cells in culture, are excellent systems for screening cytotoxicity of silver nanoparticles. These long established cell culture models and simple assays used in this study can provide useful toxicity and mechanistic information that can help to better inform safety assessments of food‐ and cosmetic‐related silver nanoparticles. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.
      PubDate: 2014-02-12T10:26:30.911359-05:
      DOI: 10.1002/jat.2994
  • Growth evaluation method by live imaging of Daphnia magna and its
           application to the estimation of an insect growth regulator
    • Authors: Akiko Suzuki; Yasuhiko Kato, Tomoaki Matsuura, Hajime Watanabe
      Abstract: The zooplankton Daphnia magna has been widely used as a test organism to assess the toxicity of chemical substances because of its important position in aquatic ecology and its ease of handling. Among the various endpoints for toxicity evaluation, growth rate is one of the most critical and many studies have been conducted. However, measurement of growth rate was time‐consuming and not an ideal endpoint in terms of screening. In this study, we demonstrated a live imaging method to monitor the growth of daphnids by area measurement. In this method, daphnid images were directly obtained from a swimming chamber and these images were processed for the evaluation of growth. The reliability of this method was confirmed by comparison with the conventional dry weight method of the same animals. The body area of daphnids using this method showed a strong correlation with the dry weight method, with R2 = 0.930. In addition, we quantified the effect of a toxicant, fenoxycarb, on the growth of the animal. Fenoxycarb concentrations of 0, 0.027, 0.27 and 2.7 µg l–1 were tested and their effects on growth were estimated by the live imaging method. In the toxicity test, the area of daphnids decreased significantly with increasing fenoxycarb concentration. These results indicate that the present live imaging method is a reliable approach for daphnid toxicity testing. This method is promising for high‐throughput Daphnia toxicity tests and real‐time individual observations. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-12T05:35:54.135943-05:
      DOI: 10.1002/jat.2988
  • Fish multigeneration test with preliminary short‐term reproduction
           assay for estrone using Japanese medaka (Oryzias latipes)
    • Authors: Ataru Nakamura; Ikumi Tamura, Hitomi Takanobu, Masumi Yamamuro, Taisen Iguchi, Norihisa Tatarazako
      Abstract: The most potent chemicals potentially causing adverse effects on fish species are estrogens in human waste. Sewage is a source of these estrogens and it is difficult to reduce. In particular, although the bioactivity of estrone is estimated to be about half of that of estradiol, multiple studies report that more than 100 ng l–1 of estrone can be detected in urban rivers, including discharges from sewage treatment works; approximately two times as high as estradiol. Few studies have been conducted to investigate the long‐term effects of estrone on wildlife; therefore, we conducted fish multigeneration test using Japanese medaka (Oryzias latipes). Medaka were exposed to estrone for 27 weeks across three generations in environmentally relevant concentrations, being 5.74, 11.4, 24.0, 47.1 and 91.4 ng l–1. No effects on reproduction were observed in the first generation; however, a decline in egg production and fertility was observed in the second generation exposed to 91.4 ng l–1 estrone, which is lower than some known environmental concentrations in urban environments. Furthermore, histopathological abnormalities were observed in the third generation exposed to both 47.1 and 91.4 ng l–1, suggesting that estrone possibly exerts severe effects on the third or later generations. However, appearances of testis–ova were observed in the second and third generation they were not consistent with actual effects on reproduction, notwithstanding the testis‐ova is regarded as the key evidence for endocrine disruption. Accordingly, we consider that qualitative measurement of abnormalities using histopathological observations is required for appropriate evaluation of endocrine disruption. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-12T05:32:04.421935-05:
      DOI: 10.1002/jat.2981
  • Microvesicle‐associated microRNA expression is altered upon
           particulate matter exposure in healthy workers and in A549 cells
    • Authors: Valentina Bollati; Laura Angelici, Giovanna Rizzo, Laura Pergoli, Federica Rota, Mirjam Hoxha, Francesco Nordio, Matteo Bonzini, Letizia Tarantini, Laura Cantone, Angela C. Pesatori, Pietro Apostoli, Andrea A. Baccarelli, Pier Alberto Bertazzi
      Abstract: Cardiovascular disease risk has been consistently linked with particulate matter (PM) exposure. Cell‐derived microvesicles (MVs) are released into plasma and transfer microRNAs (miRNAs) between tissues. MVs can be produced by the respiratory system in response to proinflammatory triggers, enter the circulatory system and remotely modify gene expression in cardiovascular tissues. However, whether PM affects MV signaling has never been investigated. In this study, we evaluated expression of microRNAs contained within plasma MVs upon PM exposure both in vivo and in vitro. In the in vivo study, we isolated plasma MVs from healthy steel plant workers before and after workplace PM exposure. We measured the expression of 88 MV‐associated miRNAs by real‐time polymerase chain reaction. To assess a possible source of the MV miRNAs identified in vivo, we measured their miRNA expression in PM‐treated A549 pulmonary cell lines in vitro. MiRNA profiling of plasma MVs showed 5.62‐ and 13.95‐fold increased expression of miR‐128 and miR‐302c, respectively, after 3 days of workplace PM exposure (P 
      PubDate: 2014-02-07T06:08:22.481934-05:
      DOI: 10.1002/jat.2987
  • Lipopolysaccharide exposure augments isoniazide‐induced liver injury
    • Authors: Yijing Su; Yun Zhang, Mi. Chen, Zhenzhou Jiang, Lixin Sun, Tao Wang, Luyong Zhang
      Abstract: Isoniazide (INH) is a classic antituberculosis drug associated with clinical idiosyncratic drug‐induced liver injury. It has been hypothesized that the interaction between a drug and modest inflammation results in a decreased threshold for drug toxicity. In this study, we tested the hypothesis that INH causes liver injury in rats when coadministered with lipopolysaccharide (LPS). Neither INH nor LPS alone caused liver injury. The coadministration of INH and LPS was associated with increases in serum and histopathological markers of liver injury. Tumour necrosis factor‐α expression was significantly increased in the coadministered group. The downregulation of the bile acid transporter, bile salt export pump, and multidrug resistance protein 2 at both mRNA and protein levels was observed. Furthermore, the level of Farnesoid X receptor, which regulates the bile salt export pump and multidrug resistance protein 2, were clearly decreased. These results indicate that the coadministration of nontoxic doses of LPS and INH causes liver injury; the disruption of biliary excretion is considered the primary inflammation‐related characteristic of INH‐induced hepatotoxicity. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-06T07:05:40.837044-05:
      DOI: 10.1002/jat.2979
  • Development of a multiparametric in vitro model of skin sensitization
    • Authors: Muriel Guyard‐Nicodème; Eloise Gerault, Marion Platteel, Olivier Peschard, Wilfried Veron, Philippe Mondon, Svinareff Pascal, Marc G. J. Feuilloley
      Abstract: Most animal experiments on cosmetics safety are prohibited and since March 2013, this obligation includes sensitization tests. However, until now there has been no validated alternative in vitro method. In this work, 400 compounds used in the cosmetic industry were selected to cover the greatest diversity of structures, biological activities and sensitizing potential. These molecules were submitted to a series of tests aimed at reproducing essential steps in sensitization and to distinguish between sensitization and irritations, i.e., transcutaneous permeation (factor A), haptenation (factor B), sensitization cytokines production (factor C) and acute toxicity (factor D). The transcutaneous diffusion was measured on human skin explants using Franz cells. Haptenation was tested in solution on human serum albumin. Sensitization cytokine production was investigated by measurement of interleukin‐18 release by keratinocytes. Acute toxicity was determined using an 3‐(4,5‐Dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide75 cell viability test. As only sufficiently stable, soluble and detectable compounds are usable, 33, 72, 68 and 68 molecules were finally tested on factors A, B, C and D, respectively, and 32 were completely screened by the four factors. The individual correlation of the four factors with the reference in vivo tests was limited but the combination of these factors led to a correlation between in vivo and in vitro assays of 81.2% and the safety of the test (risk of false negative) reached 96.8%. The techniques employed are simple and inexpensive and this model of four tests appears as a promising technique to evaluate in vitro the skin sensitization potential of unknown molecules. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-02-03T20:58:18.785567-05:
      DOI: 10.1002/jat.2986
  • A proposal to improve clarity and communication in the EU Classification
           process for chemicals for carcinogenicity and reproductive and
           developmental toxicity
    • Authors: J. E. Doe
      First page: 1068
      Abstract: There is an issue in the EU classification of substances for carcinogenicity and for reproductive or developmental toxicity which has brought difficulties to those involved in the process. The issue lies in the inability of the classification system to distinguish between carcinogens and reproductive toxicants with different levels of concern. This has its origins in the early years of toxicology when it was thought that a relatively small number of chemicals would be either carcinogens or reproductive toxicants, but this has turned out not to be the case. This can cause problems in communicating to the users of chemicals, including the public, the nature of the hazard presented by chemicals. Processes have been developed within the classification system for setting specific concentration limits which assess the degree of hazard for carcinogens and reproductive toxicants as high, medium or low. However these categories are not otherwise used in classification. It is proposed that their wider use would bring the advantages of transparency, clarity of communication, certainty of the process and would allow chemicals with a high degree of hazard to be identified and managed in an appropriate way. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-25T05:57:13.466406-05:
      DOI: 10.1002/jat.3045
  • Diisocyanates, occupational asthma and IgE antibody: implications for
           hazard characterization
    • Authors: Ian Kimber; Rebecca J. Dearman, David A. Basketter
      First page: 1073
      Abstract: Sensitization of the respiratory tract by chemicals resulting in rhinitis and asthma is an important occupational health issue. Occupational asthma is associated with significant morbidity and can be fatal. Tests for the identification and characterization of chemicals with the potential to cause sensitization of the respiratory tract are lacking. In spite of sustained interest there are no validated or widely accepted methods available, and this presents toxicologists with a considerable challenge. One important constraint on the development of appropriate testing strategies has been uncertainty and controversy about the immunological mechanisms through which chemicals may induce sensitization of the respiratory tract. By analogy with protein respiratory allergy it is legitimate to consider that IgE antibody‐dependent mechanisms may play a pivotal role. However, although many aspects of chemical respiratory allergy are consistent with reactions caused by IgE antibody, uncertainty remains because among patients with occupational asthma caused by chemical respiratory allergens there are commonly a proportion, and sometimes a significant proportion, of subjects that lack detectable IgE antibody. Here we consider the relevance of IgE antibody responses for the development of a chemical respiratory allergy to diisocyanates. A case is made that IgE antibody responses are, either directly or indirectly, closely associated with occupational asthma to the diisocyanates (and to other chemical respiratory allergens). As such the argument is advanced here that IgE antibody represents an appropriate readout for the characterization of chemical respiratory allergens, and that uncertainty about mode of action should no longer represent a hurdle in the development of suitable test methods. Copyright © 2014 John Wiley & Sons, Ltd.
      PubDate: 2014-07-25T05:48:29.612438-05:
      DOI: 10.1002/jat.3041
  • Long‐term maintenance of HepaRG cells in serum‐free conditions
           and application in a repeated dose study
    • Authors: Sebastian Klein; Daniel Mueller, Valery Schevchenko, Fozia Noor
      First page: 1078
      Abstract: Chronic repeated‐dose toxicity studies are still carried out on animals and often do not correlate with the effects in human beings mainly due to species‐specific differences in biotransformation. The human hepatoma cell line HepaRG has been used for human relevant toxicity assessment. However, HepaRG cells are commonly maintained in serum containing medium which limits their use in ’omics‘‐based toxicology. In this study, we compared the maintenance of HepaRG cells in standard serum‐supplemented and serum‐free conditions. Viability and Cytochrome P450 (CYP) activity during long‐term cultivation were assessed. Liver‐specific albumin and urea production was measured. The extracellular metabolome (amino acids, glucose, lactate and pyruvate) was measured to compare different cultivation conditions using metabolic flux analysis. Although metabolic flux analysis reveals differences in certain parts of the metabolism, e.g. production of urea, the overall metabolism of serum‐free and serum‐supplemented cultured HepaRG cells is similar. We conclude that HepaRG cells can be maintained in optimized serum‐free conditions for 30 days without viability change and with high CYP activity. We also tested the acute (24 h) and long‐term repeated‐dose (7 doses, every second day) toxicity of valproic acid. We calculated an EC50 value of 1.4 mM after repeated exposure which is close to the cmax value for valproic acid. Maintenance of HepaRG cells in serum‐free conditions opens up the opportunity for the use of these cells in human long‐term repeated‐dose hepatotoxicity studies and for application in systems toxicology. Copyright © 2013 John Wiley & Sons, Ltd.
      PubDate: 2013-09-30T09:34:10.020557-05:
      DOI: 10.1002/jat.2929
  • Identification of metabolomic biomarkers for drug‐induced acute
           kidney injury in rats
    • Authors: Takeki Uehara; Akira Horinouchi, Yuji Morikawa, Yutaka Tonomura, Keiichi Minami, Atsushi Ono, Jyoji Yamate, Hiroshi Yamada, Yasuo Ohno, Tetsuro Urushidani
      First page: 1087
      Abstract: Nephrotoxicity is a common side effect observed during both nonclinical and clinical drug development investigations. The present study aimed to identify metabolomic biomarkers that could provide early and sensitive indication of nephrotoxicity in rats. Metabolomic analyses were performed using capillary electrophoresis–time‐of‐flight mass spectrometry on rat plasma collected at 9 and 24 h after a single dose of 2‐bromoethylamine or n‐phenylanthranilic acid and at 24 h after 7 days of repeated doses of gentamicin, cyclosporine A or cisplatin. Among a total of 169 metabolites identified, 3‐methylhistidine (3‐MH), 3‐indoxyl sulfate (3‐IS) and guanidoacetate (GAA) were selected as candidate biomarkers. The biological significance and reproducibility of the observed changes were monitored over time in acute nephrotoxicity model rats treated with a single dose of cisplatin, with the glomerular filtration rate monitored by determination of creatinine clearance. Increased plasma levels of 3‐MH and 3‐IS were related to a decline in glomerular filtration due to a renal failure. In contrast, the decrease in plasma GAA, which is synthesized from arginine and glycine in the kidneys, was considered to reflect decreased production due to renal malfunction. Although definitive validation studies are required to confirm their usefulness and reliability, 3‐MH, 3‐IS and GAA may prove to be valuable plasma biomarkers for monitoring nephrotoxicity in rats. Copyright © 2013 John Wiley & Sons, Ltd.
      PubDate: 2013-09-30T09:34:05.850685-05:
      DOI: 10.1002/jat.2933
  • Prophylactic administration of non‐organophosphate cholinesterase
           inhibitors before acute exposure to organophosphates: assessment using
           terbufos sulfone
    • Authors: Dietrich E. Lorke; Syed M. Nurulain, Mohamed Y. Hasan, Kamil Kuča, Georg A. Petroianu
      First page: 1096
      Abstract: Poisoning with organophosphorus compounds (OPCs) poses a serious threat worldwide. OPC‐induced mortality can be significantly reduced by prophylactic administration of reversible acetylcholinesterase (AChE) inhibitors. The only American Food and Drug Administration (FDA)‐approved substance for such pre‐treatment (to soman exposure) is presently pyridostigmine, although its efficacy is controversial. In search for more efficacious and broad‐spectrum alternatives, we have assessed in vivo the mortality‐reducing efficacy of a group of five compounds with known AChE inhibitory activity (pyridostigmine, physostigmine, ranitidine, tacrine and K‐27), when given in equitoxic dosage (25% of LD01) 30 min before exposure to the OPC terbufos sulfone. Protection was quantified in rats by determining the relative risk of death (RR) using Cox analysis, with RR = 1 for animals given only terbufos sulfone, but no pre‐treatment. All tested AChE inhibitors reduced terbufos sulfone‐induced mortality significantly (p ≤ 0.05) as compared with the non‐treatment group (RR = 1: terbufos sulfone only). Best in vivo protection from terbufos sulfone‐induced mortality was achieved, when K‐27 was given before terbufos sulfone exposure (RR = 0.06), which was significantly (P ≤ 0.05) superior to the pre‐treatment with all other tested compounds, for example tacrine (RR = 0.21), pyridostigmine (RR = 0.28), physostigmine (RR = 0.29) and ranitidine (RR = 0.33). The differences in efficacy between tacrine, pyridostigmine, physostigmine and ranitidine were not statistically significant. Prophylactic administration of an oxime (such as K‐27) in case of imminent OPC exposure may be a viable option. Copyright © 2013 John Wiley & Sons, Ltd.
      PubDate: 2013-10-17T23:58:59.528494-05:
      DOI: 10.1002/jat.2939
  • Rapid determination of quetiapine in blood by gas
           chromatography–mass spectrometry. Application to post‐mortem
    • Authors: Olga López‐Guarnido; María Jesús Tabernero, Antonio F. Hernández, Lourdes Rodrigo, Ana M. Bermejo
      First page: 1104
      Abstract: A simple, fast and sensitive method for the determination of quetiapine in human blood has been developed and validated. The method involved a basic liquid–liquid extraction procedure and subsequent analysis by gas chromatography–mass spectrometry, previous derivatization with bis(trimethylsilyl)‐trifluoro‐acetamide and chorotrimethylsilane (99 : 1). The methods of validation included linearity with a correlation coefficient > 0.99 over the range 0.02–1 µg ml–1, intra‐ and interday precision (always 
      PubDate: 2013-10-11T06:39:37.640985-05:
      DOI: 10.1002/jat.2944
  • Borrelidin has limited anti‐cancer effects in bcl‐2
           overexpressing breast cancer and leukemia cells and reveals toxicity in
           non‐malignant breast epithelial cells
    • Authors: Diana Gafiuc; Marlene Weiß, Ioannis Mylonas, Ansgar Brüning
      First page: 1109
      Abstract: Clinically effective anti‐cancer drugs have to tread a narrow line between selective cytotoxicity on tumor cells and tolerable adverse effects against healthy tissues. This causes the failure of many potential cancer drugs in advanced clinical trials, hence signifying the importance of a comprehensive initial estimate of the cytotoxicity of prospective anti‐cancer drugs in preclinical studies. In this study, the cytotoxicity of borrelidin, a macrolide antibiotic with a high cytotoxic selectivity for proliferating endothelial cells and leukemia cells, was tested on malignant and non‐malignant breast cells. Highly metastatic breast cancer cell lines (MDA‐MB‐231 and MDA‐MB‐435) showed promising results and exhibited good sensitivity to borrelidin at low nanomolar concentrations, but borrelidin was cytotoxic to a non‐malignant breast epithelial cell line (MCF10A) as well. Furthermore, although a high sensitivity of endothelial cells (human umbilical vein endothelial cells; HUVEC) and individual leukemia cell lines (Jurkat and IM9) to borrelidin was confirmed in this study, another leukemia cell line (HL60) and an immortalized endothelial cell line (EA.hy926) displayed a significantly decreased sensitivity. Reduced sensitivity to borrelidin was associated with elevated bcl‐2 expression in these cell lines. In conclusion, the results presented show that borrelidin displays high and selective cytotoxicity against subgroups of cancer cells and endothelial cells, but, owing to its non‐specific toxicity to non‐malignant cells, its clinical application might be restricted because of likely adverse effects and limited efficacy in bcl2‐overexpressing cancer cells. Copyright © 2013 John Wiley & Sons, Ltd.
      PubDate: 2013-10-24T04:50:22.163232-05:
      DOI: 10.1002/jat.2946
  • Exposure to MnCl2 · 4H2O during development induces
           activation of microglial and perivascular macrophage populations in the
           hippocampal dentate gyrus of rats
    • Abstract: Developmental exposure to Mn caused Mn accumulation in the brain tissue and transient disruption of granule cell neurogenesis, targeting the late stage differentiation of progenitor cells in the subgranular zone of the hippocampal dentate gyrus of rats. Because neurogenesis is influenced by proinflammatory responses, this study was performed to determine whether Mn exposure causes microglial activation in the dentate hilus, a region anatomically close to the subgranular zone of the dentate gyrus. Pregnant rats were treated with dietary MnCl2 · 4H2O at 32, 160 or 800 ppm from gestational day 10 to day 21 after delivery. An immunohistochemical analysis revealed increases in Iba1+ microglia in the hilus on postnatal day 21 following exposure to MnCl2 · 4H2O in a dose‐unrelated manner at 32 and at 800 ppm and an increase in CD163+ macrophage at 800 ppm in the hilus. Real‐time reverse transcription–polymerase chain reaction analysis revealed increases in the mRNA levels of Il1α, Il6, Nos2 and Tnf after 800 ppm MnCl2 · 4H2O. These results suggest that activation of microglia and perivascular macrophages occurs in the hilus after developmental exposure to MnCl2 · 4H2O at 800 ppm, and probably involves the disruption of neurogenesis through the accumulation of Mn in the brain tissue. Copyright © 2014 John Wiley & Sons, Ltd.
  • Cylindrospermopsin induces oxidative stress and genotoxic effects in the
           fish CLC cell line
    • Abstract: Cylindrospermopsin (CYN) is a cyanotoxin detected in water reservoirs worldwide. The toxin is a potent protein synthesis inhibitor capable of adversely influencing a wide range of cell functions. While data on the prooxidative potency of CYN are inconsistent, genotoxic effects towards certain mammalian cell types have been described. However, such a potential on fish cells has not yet been investigated. Hence, the aim of the study was to elucidate the prooxidative and genotoxic impact of CYN on a common carp (Cyprinus carpio L.) leucocyte cell line (CLC). The cells were incubated with the cyanotoxin at concentrations of 0.1, 0.5 or 1 µg ml–1. After 24 h, cytotoxic activity of CYN at the highest used concentration was confirmed by decreased cell membrane integrity and inhibited cell proliferation. Additionally, CYN at 0.5 and 1 µg ml–1 increased intracellular ATP levels and decreased the reduced to oxidized glutathione ratio. Furthermore, a significant increase in the reactive oxygen species (ROS) production with concomitant changes in superoxide dismutase activity was observed after a 3.5‐h exposure of the cells to the toxin. Genotoxic activity of CYN, manifested as oxidative DNA damage and elevated number of micronuclei, was also detected in exposed cells. The obtained results indicate that CYN is able to exert a wide range of adverse effects, including oxidative stress and genotoxicity in fish leucocytes. Copyright © 2014 John Wiley & Sons, Ltd.
  • Graphene supports in vitro proliferation and osteogenic differentiation of
           goat adult mesenchymal stem cells: potential for bone tissue engineering
    • Abstract: Current treatments for bone loss injuries involve autologous and allogenic bone grafts, metal alloys and ceramics. Although these therapies have proved useful, they suffer from inherent challenges, and hence, an adequate bone replacement therapy has not yet been found. We hypothesize that graphene may be a useful nanoscaffold for mesenchymal stem cells and will promote proliferation and differentiation into bone progenitor cells. In this study, we evaluate graphene, a biocompatible inert nanomaterial, for its effect on in vitro growth and differentiation of goat adult mesenchymal stem cells. Cell proliferation and differentiation are compared between polystyrene‐coated tissue culture plates and graphene‐coated plates. Graphitic materials are cytocompatible and support cell adhesion and proliferation. Importantly, cells seeded on to oxidized graphene films undergo osteogenic differentiation in fetal bovine serum‐containing medium without the addition of any glucocorticoid or specific growth factors. These findings support graphene's potential to act as an osteoinducer and a vehicle to deliver mesenchymal stem cells, and suggest that the combination of graphene and goat mesenchymal stem cells provides a promising construct for bone tissue engineering. Copyright © 2014 John Wiley & Sons, Ltd.
  • Early chronic lead exposure reduces exploratory activity in young C57BL/6J
    • 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.
  • Development of haemostatic decontaminants for the treatment of wounds
           contaminated with chemical warfare agents. 2: Evaluation of in vitro
           topical decontamination efficacy using undamaged skin
    • Abstract: The risk of penetrating, traumatic injury occurring in a chemically contaminated environment cannot be discounted. Should a traumatic injury be contaminated with a chemical warfare (CW) agent, it is likely that standard haemostatic treatment options would be complicated by the need to decontaminate the wound milieu. Thus, there is a need to develop haemostatic products that can simultaneously arrest haemorrhage and decontaminate CW agents. The purpose of this study was to evaluate a number of candidate haemostats for efficacy as skin decontaminants against three CW agents (soman, VX and sulphur mustard) using an in vitro diffusion cell containing undamaged pig skin. One haemostatic product (WoundStat™) was shown to be as effective as the standard military decontaminants Fuller's earth and M291 for the decontamination of all three CW agents. The most effective haemostatic agents were powder‐based and use fluid absorption as a mechanism of action to sequester CW agent (akin to the decontaminant Fuller's earth). The envisaged use of haemostatic decontaminants would be to decontaminate from within wounds and from damaged skin. Therefore, WoundStat™ should be subject to further evaluation using an in vitro model of damaged skin. Copyright © 2014 Crown copyright. Journal of Applied Toxicology © 2014 John Wiley & Sons, Ltd.
  • Non‐clinical safety evaluation of single and repeated intramuscular
           administrations of MAGE‐A3 Cancer Immunotherapeutic in rabbits and
           cynomolgus monkeys
    • 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.
  • Single‐walled carbon nanotube and graphene nanodelivery of gambogic
           acid increases its cytotoxicity in breast and pancreatic cancer cells
    • Abstract: Graphene and single‐walled carbon nanotubes were used to deliver the natural low‐toxicity drug gambogic acid (GA) to breast and pancreatic cancer cells in vitro, and the effectiveness of this complex in suppressing cellular integrity was assessed. Cytotoxicity was assessed by measuring lactate dehydrogenase release, mitochondria dehydrogenase activity, mitochondrial membrane depolarization, DNA fragmentation, intracellular lipid content, and membrane permeability/caspase activity. The nanomaterials showed no toxicity at the concentrations used, and the antiproliferative effects of GA were significantly enhanced by nanodelivery. The results suggest that these complexes inhibit human breast and pancreatic cancer cells grown in vitro. This analysis represents a first step toward assessing their effectiveness in more complex, targeted, nanodelivery systems. Copyright © 2014 John Wiley & Sons, Ltd.
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