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  • Deciphering the transcription factor-microRNA-target gene regulatory network associated with graphene oxide cytotoxicity
    Nanotoxicology (IF 5.811) Pub Date : 2018-10-16
    Masoumeh Farahani, Mostafa Rezaei–Tavirani, Hakimeh Zali, Afsaneh Arefi Oskouie, Meisam Omidi, Alireza Lashay

    Graphene oxide (GO) has recently emanated as a promising material in cancer treatment. To unveil the underlying mechanisms of microRNAs (miRNAs) and potential target genes involved in GO cytotoxicity, we firstly compiled GO-related miRNAs and genes in human cancer cell lines treated with GO from public databases and published works. Besides miRNAs as post-transcriptional regulators of gene expression, transcription factors (TFs) are also the main regulators at the transcriptional level. In the following, we explored the regulatory relationships between miRNAs, target genes, and TFs. Thereafter, a gene regulatory network consisting of GO-responsive miRNAs, GO-responsive genes, and known human TFs was constructed. Then, 3-node regulatory motif types were detected in the resulting network. Among them, miRNA-FFL (feed-forward loop) was identified as a significant motif type. A total of 184 miRNA-FFLs were found and merged to generate a regulatory sub-network. Pathway analysis of the resulting sub-network highlighted adherens junction, focal adhesion, and TGFβ signaling pathways as the major pathways that previous studies demonstrate them to be the affected pathways in GO-treated cells. Functional investigations displayed that miRNAs might be involved in the control of apoptosis through disruption of cell adhesion in response to cytotoxicity. Moreover, GO-cell interactions can lead to miRNA targeting of genes (i.e. Rac1 and RhoA) involved in the cytoskeleton assembly process. These specific toxic properties support biomedical applications of GO, especially for cancer therapy.

  • Role of p53 in the chronic pulmonary immune response to tangled or rod-like multi-walled carbon nanotubes
    Nanotoxicology (IF 5.811) Pub Date : 2018-10-14
    Katherine S. Duke, Elizabeth A. Thompson, Mark D. Ihrie, Alexia J. Taylor-Just, Elizabeth A. Ash, Kelly A. Shipkowski, Jonathan R. Hall, Debra A. Tokarz, Mark F. Cesta, Ann F. Hubbs, Dale W. Porter, Linda M. Sargent, James C. Bonner

    The fiber-like shape of multi-walled carbon nanotubes (MWCNTs) is reminiscent of asbestos, suggesting they pose similar health hazards when inhaled, including pulmonary fibrosis and mesothelioma. Mice deficient in the tumor suppressor p53 are susceptible to carcinogenesis. However, the chronic pathologic effect of MWCNTs delivered to the lungs of p53 heterozygous (p53+/−) mice has not been investigated. We hypothesized that p53+/− mice would be susceptible to lung tumor development after exposure to either tangled (t-) or rod-like (r-) MWCNTs. Wild-type (p53+/+) or p53+/− mice were exposed to MWCNTs (1 mg/kg) via oropharyngeal aspiration weekly over four consecutive weeks and evaluated for cellular and pathologic outcomes 11-months post-initial exposure. No lung or pleural tumors were observed in p53+/+ or p53+/− mice exposed to either t- or rMWCNTs. In comparison to tMWCNTs, the rMWCNTs induced the formation of larger granulomas, a greater number of lymphoid aggregates and greater epithelial cell hyperplasia in terminal bronchioles in both p53+/− and p53+/+ mice. A constitutively larger area of CD45R+/CD3+ lymphoid tissue was observed in p53+/− mice compared to p53+/+ mice. Importantly, p53+/− mice had larger granulomas induced by rMWCNTs as compared to p53+/+ mice. These findings indicate that a combination of p53 deficiency and physicochemical characteristics including nanotube geometry are factors in susceptibility to MWCNT-induced lymphoid infiltration and granuloma formation.

  • Uptake and molecular impact of aluminum-containing nanomaterials on human intestinal caco-2 cells
    Nanotoxicology (IF 5.811) Pub Date : 2018-10-14
    Holger Sieg, Caroline Braeuning, Birgitta Maria Kunz, Hannes Daher, Claudia Kästner, Benjamin-Christoph Krause, Thomas Meyer, Pégah Jalili, Kevin Hogeveen, Linda Böhmert, Dajana Lichtenstein, Agnès Burel, Soizic Chevance, Harald Jungnickel, Jutta Tentschert, Peter Laux, Albert Braeuning, Fabienne Gauffre, Valérie Fessard, Jan Meijer, Irina Estrela-Lopis, Andreas F. Thünemann, Andreas Luch, Alfonso Lampen

    Aluminum (Al) is one of the most common elements in the earth crust and increasingly used in food, consumer products and packaging. Its hazard potential for humans is still not completely understood. Besides the metallic form, Al also exists as mineral, including the insoluble oxide, and in soluble ionic forms. Representatives of these three species, namely a metallic and an oxidic species of Al-containing nanoparticles and soluble aluminum chloride, were applied to human intestinal cell lines as models for the intestinal barrier. We characterized physicochemical particle parameters, protein corona composition, ion release and cellular uptake. Different in vitro assays were performed to determine potential effects and molecular modes of action related to the individual chemical species. For a deeper insight into signaling processes, microarray transcriptome analyses followed by bioinformatic data analysis were employed. The particulate Al species showed different solubility in biological media. Metallic Al nanoparticles released more ions than Al2O3 nanoparticles, while AlCl3 showed a mixture of dissolved and agglomerated particulate entities in biological media. The protein corona composition differed between both nanoparticle species. Cellular uptake, investigated in transwell experiments, occurred predominantly in particulate form, whereas ionic Al was not taken up by intestinal cell lines. Transcellular transport was not observed. None of the Al species showed cytotoxic effects up to 200 µg Al/mL. The transcriptome analysis indicated mainly effects on oxidative stress pathways, xenobiotic metabolism and metal homeostasis. We have shown for the first time that intestinal cellular uptake of Al occurs preferably in the particle form, while toxicological effects appear to be ion-related.

  • Autophagy-dependent release of zinc ions is critical for acute lung injury triggered by zinc oxide nanoparticles
    Nanotoxicology (IF 5.811) Pub Date : 2018-10-14
    Xuejun Jiang, Qianghu Tang, Jun Zhang, Hong Wang, Lulu Bai, Pan Meng, Xia Qin, Ge Xu, Diptiman D. Bose, Bin Wang, Chengzhi Chen, Zhen Zou

    Pulmonary exposure to zinc oxide nanoparticles (ZnONPs) could cause acute lung injury (ALI), but the underlying molecular mechanism remains unclear. Herein, we established a ZnONPs-induced ALI mouse model, characterized by the histopathological changes (edema and infiltration of inflammatory cells in lung tissues), and the elevation of total protein and cytokine interleukin-6 in bronchoalveolar lavage fluid in time- and dose-dependent manners. This model also exhibited features like the disturbance of redox-state (reduced of glutathione to glutathione disulfide ratio, elevation of heme oxygenase-1 and superoxide dismutase 2), the decrease of adenosine triphosphate synthesis and the release of zinc ions in the lung tissues. Interestingly, we found that ZnONPs exposure caused the accumulation of autophagic vacuoles and the elevation of microtubule-associated proteins 1A/1B light chain (LC)3B-II and p62, indicating the impairment of autophagic flux. Our data indicated that the above process might be regulated by the activation of AMP-activated protein kinase but not the mammalian target of rapamycin pathway. The association between ZnONPs-induced ALI and autophagy was further verified by a classical autophagy inhibitor, 3-methyladenine (3-MA). 3-MA administration reduced the accumulation of autophagic vacuoles, the expression of LC3B-II and p62, followed by a significant attenuation of histopathological changes, inflammation, and oxidative stress. More importantly, 3-MA could directly decrease the release of zinc ions in lung tissues. Taken together, our study provides the evidence that ZnONPs-induced pulmonary toxicity is autophagy-dependent, suggests that limiting the release of zinc ions by inhibiting autophagy could be a feasible strategy for the prevention of ZnONPs-associated pulmonary toxicity.

  • Comparative analysis of nanosystems’ effects on human endothelial and monocytic cell functions
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-28
    Jasmin Matuszak, Philipp Dörfler, Stefan Lyer, Harald Unterweger, Maya Juenet, Cédric Chauvierre, Amr Alaarg, Danielle Franke, Gunter Almer, Isabelle Texier, Josbert M. Metselaar, Ruth Prassl, Christoph Alexiou, Harald Mangge, Didier Letourneur, Iwona Cicha

    The objective of our work was to investigate the effects of different types of nanoparticles on endothelial (HUVEC) and monocytic cell functions. We prepared and tested 14 different nanosystems comprising liposomes, lipid nanoparticles, polymer, and iron oxide nanoparticles. Some of the tested nanosystems contained targeting, therapeutic, or contrast agent(s). The effect of particles (0–400 µg/mL) on endothelial-monocytic cell interactions in response to TNF-α was investigated using an arterial bifurcation model and dynamic monocyte adhesion assay. Spontaneous HUVEC migration (0–100 µg/mL nanoparticles) and chemotaxis of monocytic cells towards MCP-1 in presence of particles (0–400 µg/mL) were determined using a barrier assay and a modified Boyden chamber assay, respectively. Lipid nanoparticles dose-dependently reduced monocytic cell chemotaxis and adhesion to activated HUVECs. Liposomal nanoparticles had little effect on cell migration, but one formulation induced monocytic cell recruitment by HUVECs under non-uniform shear stress by about 50%. Fucoidan-coated polymer nanoparticles (25–50 µg/mL) inhibited HUVEC migration and monocytic cell chemotaxis, and had a suppressive effect on monocytic cell recruitment under non-uniform shear stress. No significant effects of iron oxide nanoparticles on monocytic cell recruitment were observed except lauric acid and human albumin-coated particles which increased endothelial-monocytic interactions by 60–70%. Some of the iron oxide nanoparticles inhibited HUVEC migration and monocytic cell chemotaxis. These nanoparticle-induced effects are of importance for vascular cell biology and function and must be considered before the potential clinical use of some of the analyzed nanosystems in cardiovascular applications.

  • Activated iRhom2 drives prolonged PM2.5 exposure-triggered renal injury in Nrf2-defective mice
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-26
    Min-Xuan Xu, Yu-Ting Qin, Chen-Xu Ge, Ting-Ting Gu, De-Shuai Lou, Qiang Li, Lin-Feng Hu, Yuan-Yuan Li, Wei-Wei Yang, Jun Tan

    Research suggests that particulate matter (PM2.5) is a predisposing factor for metabolic syndrome-related systemic inflammation and oxidative stress injury. TNF-α as a major pro-inflammatory cytokine was confirmed to participate in various diseases. Inactive rhomboid protein 2 (iRhom2) was recently determined as a necessary regulator for shedding of TNF-α in immune cells. Importantly, kidney-resident macrophages are critical to inflammation-associated chronic renal injury. Podocyte injury can be induced by stimulants and give rise to nephritis, but how iRhom2 contributes to PM2.5-induced renal injury is unclear. Thus, we studied whether PM2.5 causes renal injury and characterized iRhom2 with respect to TNF-α release in mice macrophages and renal tissues in long-term PM2.5-exposed mouse models. After long-term PM2.5 exposures, renal injury was confirmed via inflammatory cytokine, chemokine expression, and reduced antioxidant activity. Patients with kidney-related diseases had increased TNF-α, which may contribute to renal injury. We observed up-regulation of serum creatinine, serum urea nitrogen, kidney injury molecule 1, uric acid, TNF-α, MDA, H2O2, and O2– in PM2.5-treated mice, which was greater than that found in Nrf2−/− mice. Meanwhile, increases in metabolic disorder-associated indicators were involved in PM2.5-induced nephritis. In vitro, kidney-resident macrophages were observed to be critical to renal inflammatory infiltration and function loss via regulation of iRhom2/TACE/TNF-α signaling, and suppression of Nrf2-associated anti-oxidant response. PM2.5 exposure led to renal injury partly by inflammation-mediated podocyte injury. Reduced SOD1, SOD2, Nrf2 activation, and increased XO, NF-κB activity, TACE, iNOS, IL-1β, TNF-α, IL-6, MIP-1α, Emr-1, MCP-1, and Cxcr4, were also noted. Long-term PM2.5 exposure causes chronic renal injury by up-regulation of iRhom2/TACE/TNF-α axis in kidney-resident macrophages. Overexpression of TNF-α derived from macrophages causes podocyte injury and kidney function loss. Thus, PM2.5 toxicities are related to exposure duration and iRhom2 may be a potential therapeutic renal target.

  • Human nasal mucosal C-reactive protein responses after inhalation of ultrafine welding fume particles: positive correlation to systemic C-reactive protein responses
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-26
    R. Baumann, P. Brand, A. Chaker, A. Markert, I. Rack, S. Davatgarbenam, S. Joraslafsky, B. Gerhards, T. Kraus, M. Gube

    Exposures to occupationally relevant ultrafine, zinc- and copper-containing welding fumes cause inflammatory responses involving systemic IL-6, C-reactive protein (CRP) and serum amyloid A (SAA), all associated with elevated risk of cardiovascular events. We investigated whether the systemic response is preceded by nasal inflammatory reactions. Fifteen nonsmoking male subjects were exposed for 6 h under controlled conditions to zinc-/copper-containing welding fumes (at 2.5 mg/m3) or ambient air control in a randomized order. Nasal secretions were collected before and at 1, 3, 6, 10, and 29 h after exposure. Nasal levels of selected biomarkers were determined by electrochemiluminescent assays and related to their systemic levels. Nasal interferon-γ (IFN-γ) peaked significantly 1 h after start of exposure compared to baseline. Nasal CRP as well as SAA increased significantly at 10 and 29 h compared to baseline. Receiver operating characteristic (ROC) curve analysis for differentiating welding fume from control exposure was performed: The highest area under ROC curve (AUC) values were found for the CRP increases (10, 29 h versus 0 h): AUC = 0.83, and for IFN-γ increases (1 h versus 0 h): AUC = 0.92. Nasal and systemic changes of CRP at 29 h revealed a strong correlation (Spearman rank test: increases compared to baseline: r = 0.815, p = 0.0022; absolute levels: r = 0.9, p = 0.0002). In conclusion, short-term exposure to a zinc- and copper-containing welding fume causes significant increases of inflammatory mediators in nasal mucosal lining fluid. Therefore, measurement of nasal inflammatory mediators may provide a useful means for occupational surveillance of workers exposed to ultrafine metal fume particles.

  • Systematic in vivo study of NiO nanowires and nanospheres: biodegradation, uptake and biological impacts
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-25
    Mohamed Alaraby, Alba Hernández, Ricard Marcos

    High aspect ratio nanomaterials (NM) have a promising future in medicine and industry as a unique category of NM. Consequently, it is important to evaluate their potential biological side-effects before crediting their use. To understand the mechanisms of degradation, internalisation, and interaction with different biological targets, we used the in vivo model Drosophila melanogaster to obtain a systematic and complete study on high aspect ratio Ni nanowires (NiNW), compared with low aspect ratio Ni nanospheres (NiNS), and NiSO4 as a model of agent releasing nickel ions. The distinguished shape of nanowires showed changes in their characteristics after oral administration until they reached the intestinal lumen, where their diameter decreased significantly. For the first time, we confirmed the internalization of needle-shaped materials via perforation of the intestinal barrier. Moreover, the results showed that D. melanogaster is a valid and effective tool in studies related to magnetic resonance imaging (MRI). Additionally, NiNM induced DNA damage and molecular changes at the gene expression level, in association with increase in oxidative stress levels. Notably, the observed negative effects were related to nickel as a metal rather than to its shape, since the effects induced by the three Ni forms were notably similar. In addition, independent of their form, Ni compounds did not induce toxic or mutagenic impacts. Our Drosophila model can be used to understand different phenomena related to high aspect ratio NM exposure, such as degradation, internalization and interaction with different targets.

  • Differential toxicity of processed and non-processed states of CoCrMo degradation products generated from a hip simulator on neural cells
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-25
    Divya Rani Bijukumar, Abhijith Segu, YongChao Mou, Reza Ghodsi, Tolou Shokufhar, Mark Barba, Xue-Jun Li, Mathew Thoppil Mathew

    Physico-chemical characteristics of the CoCrMo degradation products have played an important role in cytotoxicity and clinical complications on the orthopedic patients who have metal implants. Previous studies have limited reflection on the physicochemical characteristics of the degradation products generated in vivo, which are very different from individual metal particles and/or ions obtained from different commercial sources. In this study, we aimed to understand the differences in toxicity induced by the degradation products in as-synthesized form as well as those obtained after post-processing. The degradation products were generated using a hip-simulator by maintaining physiological conditions closer to in vivo and separated into two batches, one with processing by washing and drying called processed degradation products (PDP) and another batch as ‘as-synthesized’ degradation product (DP). We studied the dose-dependent toxicity response by neural cells derived from induced pluripotent stem cells. The results of the study show that as-synthesized DPs are more toxic to neural cells even at lower concentrations studied with evident low TC50 (1–5 μg/ml) concentrations compared to PDP (25 μg/ml). Flow cytometric analysis showed a significant (p<.01) increase in uptake of the particles after 24 h and corresponding ROS production in DP-treated cells. RT-PCR analysis of oxidative specific gene expression showed, elevated mRNA levels of NADPH oxidase-1, nuclear transcription factor, superoxide dismutase-2 and glutaredoxin-2 in DP-treated cells after 6 h. The results of the study provided a clear evidence of the differential response of neural cells on the degradation products as a function of concentrations and their chemical nature.

  • Evaluation of toxicological and teratogenic effects of carbosilane glucose glycodendrimers in zebrafish embryos and model rodent cell lines
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-05
    Michaela Liegertová, Dominika Wrobel, Regina Herma, Monika Müllerová, Lucie Červenková Šťastná, Petra Cuřínová, Tomáš Strašák, Marek Malý, Jan Čermák, Jiří Smejkal, Marcel Štofik, Jan Maly

    Glycodendrimers (Glyco-DDMs) represent a rapidly growing class of nanoparticles with promising properties for biomedical applications but concerns regarding the impact on human health and environment are still justified. Here we report, for the first time, the comparative study of in vivo developmental toxicity of carbosilane Glyco-DDMs and their cytotoxicity in vitro. Carbosilane Glyco-DDMs (generation 1–3) containing 4, 8, and 16 β-d-glucopyranosyl units at the periphery (DDM1Glu, DDM2Glu, and DDM3Glu) were synthesized and characterized by 1H, 13C and 29Si NMR, mass spectrometry, dynamic light scattering, atomic force microscopy (AFM), and computer modeling. In vitro cytotoxicity assay (MTT) of DDM1–3Glu was performed on three different rodent cell lines (Cricetulus griseus) – B14 (ATCC, CCL-14.1), BRL 3A (ATCC, CRL-1442), and NRK 52E (ATCC, CRL-1571). Overall, very low cytotoxicity was observed with calculated IC50 in mM range with slight difference between each cell line and DDM generation investigated. Modified fish embryo test (FET) was further used for DDM3Glu developmental toxicity testing on zebrafish (Danio rerio) embryos. While seemingly harmless to intact embryos, adverse effects of DDMs on the embryonic development become evident after chorion removal (LD50=2.78 µM at 96 hpe). We summarized that the modified FET test showed a two to three orders of magnitude difference between the in vitro cytotoxicity and in vivo developmental toxicity of DDM3Glu. While, in general, the Glyco-DDMs show great promises as efficient vectors in targeted drug delivery or as therapeutic molecules itself, we suggest that their developmental toxicity should be thoroughly investigated to exclude safety risks associated with their potential biomedical use.

  • Mechanism of long-term toxicity of CuO NPs to microalgae
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-05
    Xingkai Che, Ruirui Ding, Yuting Li, Zishan Zhang, Huiyuan Gao, Wei Wang

    Little is known regarding the detailed mechanism of CuO NPs’ toxicity to microalgal primary metabolism pathway. Photosynthesis and respiration are the most important primary metabolism and the main sources of production of reactive oxygen species (ROS), but the effect of CuO NPs on both of them has not been systematically studied to date. Our research demonstrated that long-term treatment with CuO NPs significantly inhibited activities of photosynthesis and respiration in microalgae, and the photosynthesis was more sensitive to the toxicity of CuO NPs than respiration. CuO NPs could be absorbed by microalgae and be converted into Cu2O NPs concentrated in chloroplast. The internalized Cu, regardless of whether the exposure was Cu2+ or CuO NPs had the same capacity to damage chloroplast structure. The result also shows that the oxygen-evolving complex (OEC) in the photosynthetic electron transport chain was the most sensitive site to CuO NPs and Cu2+-treated microalgae had the same damage site as that of CuO NPs, which may be related to the Mn cluster that is dissociated by Cu ions released from CuO NPs. The damage of OEC inhibited photosynthetic electron transport to increase excess excited energy, which caused the accumulation of ROS in chloroplast. The accumulation of ROS damaged the structure of cell membrane and aggravated the PSII photoinhibition, further decreasing the efficiency of light energy utilization. In conclusion, the Cu ionic toxicity of photosynthetic apparatus by CuO NPs resulted in the carbon starvation and the accumulation of ROS to inhibit the growth of microalgae.

  • Preclinical hazard evaluation strategy for nanomedicines
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-05
    Stefan Siegrist, Emre Cörek, Pascal Detampel, Jenny Sandström, Peter Wick, Jörg Huwyler

    The increasing nanomedicine usage has raised concerns about their possible impact on human health. Present evaluation strategies for nanomaterials rely on a case-by-case hazard assessment. They take into account material properties, biological interactions, and toxicological responses. Authorities have also emphasized that exposure route and intended use should be considered in the safety assessment of nanotherapeutics. In contrast to an individual assessment of nanomaterial hazards, we propose in the present work a novel and unique evaluation strategy designed to uncover potential adverse effects of such materials. We specifically focus on spherical engineered nanoparticles used as parenterally administered nanomedicines. Standardized assay protocols from the US Nanotechnology Characterization Laboratory as well as the EU Nanomedicine Characterisation Laboratory can be used for experimental data generation. We focus on both cellular uptake and intracellular persistence as main indicators for nanoparticle hazard potentials. Based on existing regulatory specifications defined by authorities such as the European Medicines Agency and the United States Food and Drug Administration, we provide a robust framework for application-oriented classification paired with intuitive decision making. The Hazard Evaluation Strategy (HES) for injectable nanoparticles is a three-tiered concept covering physicochemical characterization, nanoparticle (bio)interactions, and hazard assessment. It is cost-effective and can assist in the design and optimization of nanoparticles intended for therapeutic use. Furthermore, this concept is designed to be adaptable for alternative exposure and application scenarios. To the knowledge of the authors, the HES is unique in its methodology based on exclusion criteria. It is the first hazard evaluation strategy designed for nanotherapeutics.

  • A nanoinformatics decision support tool for the virtual screening of gold nanoparticle cellular association using protein corona fingerprints
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-05
    Antreas Afantitis, Georgia Melagraki, Andreas Tsoumanis, Eugenia Valsami-Jones, Iseult Lynch

    The increasing use of nanoparticles (NPs) in a wide range of consumer and industrial applications has necessitated significant effort to address the challenge of characterizing and quantifying the underlying nanostructure – biological response relationships to ensure that these novel materials can be exploited responsibly and safely. Such efforts demand reliable experimental data not only in terms of the biological dose-response, but also regarding the physicochemical properties of the NPs and their interaction with the biological environment. The latter has not been extensively studied, as a large surface to bind biological macromolecules is a unique feature of NPs that is not relevant for chemicals or pharmaceuticals, and thus only limited data have been reported in the literature quantifying the protein corona formed when NPs interact with a biological medium and linking this with NP cellular association/uptake. In this work we report the development of a predictive model for the assessment of the biological response (cellular association, which can include both internalized NPs and those attached to the cell surface) of surface-modified gold NPs, based on their physicochemical properties and protein corona fingerprints, utilizing a dataset of 105 unique NPs. Cellular association was chosen as the end-point for the original experimental study due to its relevance to inflammatory responses, biodistribution, and toxicity in vivo. The validated predictive model is freely available online through the Enalos Cloud Platform (http://enalos.insilicotox.com/NanoProteinCorona/) to be used as part of a regulatory or NP safe-by-design decision support system. This online tool will allow the virtual screening of NPs, based on a list of the significant NP descriptors, identifying those NPs that would warrant further toxicity testing on the basis of predicted NP cellular association.

  • Insights into possibilities for grouping and read-across for nanomaterials in EU chemicals legislation
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-05
    A. Mech, K. Rasmussen, P. Jantunen, L. Aicher, M. Alessandrelli, U. Bernauer, E. A. J. Bleeker, J. Bouillard, P. Di Prospero Fanghella, R. Draisci, M. Dusinska, G. Encheva, G. Flament, A. Haase, Y. Handzhiyski, F. Herzberg, J. Huwyler, N. R. Jacobsen, V. Jeliazkov, N. Jeliazkova, P. Nymark, R. Grafström, A. G. Oomen, M. L. Polci, C. Riebeling, J. Sandström, B. Shivachev, S. Stateva, S. Tanasescu, R. Tsekovska, H. Wallin, M. F. Wilks, S. Zellmer, M. D. Apostolova

    This paper presents a comprehensive review of European Union (EU) legislation addressing the safety of chemical substances, and possibilities within each piece of legislation for applying grouping and read-across approaches for the assessment of nanomaterials (NMs). Hence, this review considers both the overarching regulation of chemical substances under REACH (Regulation (EC) No 1907/2006 on registration, evaluation, authorization, and restriction of chemicals) and CLP (Regulation (EC) No 1272/2008 on classification, labeling and packaging of substances and mixtures) and the sector-specific pieces of legislation for cosmetic, plant protection and biocidal products, and legislation addressing food, novel food, and food contact materials. The relevant supporting documents (e.g. guidance documents) regarding each piece of legislation were identified and reviewed, considering the relevant technical and scientific literature. Prospective regulatory needs for implementing grouping in the assessment of NMs were identified, and the question whether each particular piece of legislation permits the use of grouping and read-across to address information gaps was answered.

  • Grouping of nanomaterials to read-across hazard endpoints: a review
    Nanotoxicology (IF 5.811) Pub Date : 2018-09-05
    L. Lamon, K. Aschberger, D. Asturiol, A. Richarz, A. Worth

    The use of non-testing strategies like read-across in the hazard assessment of chemicals and nanomaterials (NMs) is deemed essential to perform the safety assessment of all NMs in due time and at lower costs. The identification of physicochemical (PC) properties affecting the hazard potential of NMs is crucial, as it could enable to predict impacts from similar NMs and outcomes of similar assays, reducing the need for experimental (and in particular animal) testing. This manuscript presents a review of approaches and available case studies on the grouping of NMs to read-across hazard endpoints. We include in this review grouping frameworks aimed at identifying hazard classes depending on PC properties, hazard classification modules in control banding (CB) approaches, and computational methods that can be used for grouping for read-across. The existing frameworks and case studies are systematically reported. Relevant nanospecific PC properties taken into account in the reviewed frameworks to support grouping are shape and surface properties (surface chemistry or reactivity) and hazard classes are identified on the basis of biopersistence, morphology, reactivity, and solubility.

  • Biodistribution and biocompatibility of passion fruit-like nano-architectures in zebrafish
    Nanotoxicology (IF 5.811) Pub Date : 2018-08-22
    Marta d’Amora, Domenico Cassano, Salvador Pocoví-Martínez, Silvia Giordani, Valerio Voliani

    Passion fruit-like nano-architectures (NAs) are all-in-one platforms of increasing interest for the translation of metal nanoparticles into clinics. NAs are nature-inspired disassembling inorganic theranostics, which jointly combine most of the appealing behaviors of noble metal nanoparticles with their potential organism excretion. Despite their unique and promising properties, NAs in vivo interactions and potential adverse effects have not yet been investigated. In this study, we employ zebrafish (Danio Rerio) to assess the development toxicity of NAs as well as their uptake and bioaccumulation at different stages of growth. The evaluation of multiple endpoints related to the toxicity clearly indicates that NAs do not induce mortality, developmental defects, or alterations on the hatching rate and behavior of zebrafish. Moreover, the analysis of nanostructures uptake and biodistribution demonstrates that NAs are successfully internalized and present a specific localization. Overall, our results demonstrate that NAs are able to pass through the embryos chorion and accumulate in specific tissues, exhibiting an impressive biocompatibility.

  • Disaggregation of gold nanoparticles by Daphnia magna
    Nanotoxicology (IF 5.811) Pub Date : 2018-07-27
    Karin Mattsson, Ruben Aguilar, Oscar Torstensson, Diana Perry, Katja Bernfur, Sara Linse, Lars-Anders Hansson, Karin S. Åkerfeldt, Tommy Cedervall

    The use of manufactured nanomaterials is rapidly increasing, while our understanding of the consequences of releasing these materials into the environment is still limited and many questions remain, for example, how do nanoparticles affect living organisms in the wild? How do organisms adapt and protect themselves from exposure to foreign materials? How does the environment affect the performance of nanoparticles, including their surface properties? In an effort to address these crucial questions, our main aim has been to probe the effects of aquatic organisms on nanoparticle aggregation. We have, therefore, carried out a systematic study with the purpose to disentangle the effects of the freshwater zooplankter, Daphnia magna, on the surface properties, stability, and aggregation properties of gold (Au) nanoparticles under different aqueous conditions as well as identified the proteins bound to the nanoparticle surface. We show that Au nanoparticles aggregate in pure tap water, but to a lesser extent in water that either contains Daphnia or has been pre-conditioned with Daphnia. Moreover, we show that proteins generated by Daphnia bind to the Au nanoparticles and create a modified surface that renders them less prone to aggregation. We conclude that the surrounding milieu, as well as the surface properties of the original Au particles, are important factors in determining how the nanoparticles are affected by biological metabolism. In a broader context, our results show how nanoparticles released into a natural ecosystem become chemically and physically altered through the dynamic interactions between particles and organisms, either through biological metabolism or through the interactions with biomolecules excreted by organisms into the environment.

  • Gold nanoparticles ingested by oyster larvae are internalized by cells through an alimentary endocytic pathway
    Nanotoxicology (IF 5.811) Pub Date : 2018-07-11
    Seta Noventa, Christian Hacker, Ana Correia, Claudia Drago, Tamara Galloway

    The biological fate of nanoparticles (NPs) taken up by organisms from their environment is a crucial issue for assessing ecological hazard. Despite its importance, it has scarcely been addressed due to the technical difficulties of doing so in whole organism in vivo studies. Here, by using transmission electron microscopy and energy dispersive X-ray spectroscopy (TEM-EDS), we describe the key aspects that characterize the interaction between an aquatic organism of global ecological and economic importance, the early larval stage of the Japanese oyster (Crassostrea gigas), and model gold NPs dispersed in their environment. The small size of the model organism allowed for a high-throughput visualization of the subcellular distribution of NPs, providing a comprehensive and robust picture of the route of uptake, mechanism of cellular permeation, and the pathways of clearance counterbalancing bioaccumulation. We show that NPs are ingested by larvae and penetrate cells through alimentary pinocytic/phagocytic mechanisms. They undergo intracellular digestion and storage inside residual bodies, before excretion with feces or translocation to phagocytic coelomocytes of the visceral cavity for potential extrusion or further translocation. Our mechanistically-supported findings highlight the potential of oyster larvae and other organisms which feature intracellular digestion processes to be exposed to man-made NPs and thus any risks associated with their inherent toxicity.

  • Antigenotoxic potential of boron nitride nanotubes
    Nanotoxicology (IF 5.811) Pub Date : 2018-06-28
    Eşref Demir, Ricard Marcos

    Boron and boron nitride nanotubes (BNNTs) are increasingly used in different industrial fields and, potentially, in some biomedical areas. As occurs with other nanomaterials (NMs), to increase our knowledge on their potential health hazards is a priority. Although in vitro approaches are a routine in getting biological information on the biological effects of NMs, the use of simple in vivo model organisms is receiving an increased interest. In this context, Drosophila melanogaster is widely used as a eukaryotic model for the study of the potential harmful effects associated with various agents, including NMs. The aim of this study is to provide new data on the potential antioxidant/antigenotoxic properties of boron and boron nitride nanotubes (BNNTs), as well as on other biological end-points. Our results show changes in the expression of genes involved in the antioxidant defense (CAT and SOD), and in those rel0061ted to the integrity of the intestinal barrier (Duox, Hml, Muc68D, and PPO2), at the highest exposure doses (5, 10 mM). However, non-relevant toxic or genotoxic effects were observed. Interestingly, BNNTs and boron significantly reduced the genotoxic effect of potassium dichromate (PDC), and the intracellular levels of reactive oxygen species (ROS). This suggest that the observed effects can be linked to the antioxidant properties of BNNTs and boron. This is the first study reporting antigenotoxicity/genotoxicity, and gene expression data, in the somatic cells of D. melanogaster larvae for BNNTs.

  • Cationic polystyrene nanoparticle and the sea urchin immune system: biocorona formation, cell toxicity, and multixenobiotic resistance phenotype
    Nanotoxicology (IF 5.811) Pub Date : 2018-06-27
    L. F. Marques-Santos, G. Grassi, E. Bergami, C. Faleri, T. Balbi, A. Salis, G. Damonte, L. Canesi, I. Corsi

    In order to assess the impact of nanoplastics on marine species, polystyrene nanoparticles (PS NPs) have been largely used as model particles. Here we studied the effects of 50 nm amino-modified PS-NH2 on Mediterranean sea urchin Paracentrotus lividus immune system cells (coelomocytes) in the presence of celomic fluid (CF) and at different NP concentrations (1, 5, 10, and 25 μg mL−1) and experimental conditions (absence or presence of EDTA). PS-NH2 acquired a protein corona once incubated with CF, dominated by the toposome precursor protein (TPP). In short-term cultures, a significant concentration- and time-dependent decrease in lysosomal membrane stability and apoptotic-like nuclear alterations were observed in phagocytes upon exposure to PS-NH2 (10 and 25 µg mL−1) in CF but they resulted abolished in the presence of EDTA confirming the role of TPP in triggering PS-NH2-coelomocytes interaction and toxicity. PS-NH2 did not alter MXR phenotype but the observed dose-dependent decrease in calcein accumulation suggests the ability of PS-NH2 to affect pump’s efflux activity. Overall results encourage additional studies on positively charged nanoplastics, since the observed effects on sea urchin coelomocytes as well as the TPP corona formation might represent a first step for addressing their impact on sensitive marine species.

  • Silver nanoparticles inhibit neural induction in human induced pluripotent stem cells
    Nanotoxicology (IF 5.811) Pub Date : 2018-06-14
    Shigeru Yamada, Daiju Yamazaki, Yasunari Kanda

    Silver nanoparticles (AgNPs) have been widely used as consumer products due to their antibacterial activities. Despite their extensive use, AgNPs have been reported to cause various types of cytotoxicity, including neurotoxicity. However, the potential action of AgNPs on early fetal development has not been elucidated. This study determined the effects of AgNPs on neural induction in human induced pluripotent stem cells (iPSCs), used as a model for human fetal stage development. It was observed that exposure to AgNPs reduced the expression of several neural differentiation marker genes, including OTX2, an early biomarker for neurogenesis in iPSCs. Since neural differentiation requires ATP as a source of energy, the intracellular ATP content was also measured. It was observed that AgNPs decreased intracellular ATP levels in iPSCs. Since AgNPs suppressed energy production, a critical mitochondrial function, the effects of AgNPs on mitochondrial dynamics were further studied. The results revealed that AgNPs induced mitochondrial fragmentation and reduced the level of mitochondrial fusion protein mitofusin 1 (Mfn1). Previously, we reported that knockdown of Mfn1 in iPSCs inhibited neural induction via OTX2 downregulation. This suggested that AgNPs could induce cytotoxicity, including neurodevelopmental toxicity, via Mfn1-mediated mitochondrial dysfunction in iPSCs. Thus, mitochondrial function in iPSCs can be used for assessing the cytotoxic effects associated with nanomaterials, including AgNPs.

  • Investigating oxidation state-induced toxicity of PEGylated graphene oxide in ocular tissue using gene expression profiles
    Nanotoxicology (IF 5.811) Pub Date : 2018-06-09
    Wei Wu, Liang Yan, Siyu Chen, Qiyou Li, Zhanjun Gu, Haiwei Xu, Zheng Qin Yin

    Graphene and its derivatives are widely used for a variety of industrial, biomedical, and environmental applications. However, the potential harm caused by exposure of the eyes to graphene-based nanomaterials is scarce. Given the potential for these materials to be used in multiple applications, there is a pressing need to evaluate their ocular toxicity, and understand the relationships between their physico-chemical properties and the resulting toxicity. In this study, the toxicity of PEGylated graphene oxide (PEG-GO) with differing oxidation levels and/or surface charges (positive, negative and neutral charge) was evaluated using two in-vitro models of the eye: primary human corneal epithelial cells and human retinal capillary endothelial cells. The results showed that oxidation level, but not surface charge, had a pivotal effect on the toxicity of graphene-based nanomaterials. Typically, PEG-GO sample with a higher oxidation level caused more serious cytotoxicity than those with a lower oxidation level. Furthermore, by analysis of global gene expression profiles, we found that the foremost cellular response to PEG-GO sample with a high oxidation level was the oxidative stress response. Next, via exploring the underlying molecular mechanism of oxidative stress-induced cytotoxicity, we showed that PEG-GO sample with a high degree of oxidation induced reactive oxygen species (ROS) via NDUFB9-mediated biological pathway. This work has significant implications for design of safe graphene-based nanomaterials for biomedical applications.

  • Genotoxicity induced by metal oxide nanoparticles: a weight of evidence study and effect of particle surface and electronic properties
    Nanotoxicology (IF 5.811) Pub Date : 2018-06-09
    Azadi Golbamaki, Nazanin Golbamaki, Natalia Sizochenko, Bakhtiyor Rasulev, Jerzy Leszczynski, Emilio Benfenati

    The genetic toxicology of nanomaterials is a crucial toxicology issue and one of the least investigated topics. Substantially, the genotoxicity of metal oxide nanomaterials’ data is resulting from in vitro comet assay. Current contributions to the genotoxicity data assessed by the comet assay provide a case-by-case evaluation of different types of metal oxides. The existing inconsistency in the literature regarding the genotoxicity testing data requires intelligent assessment strategies, such as weight of evidence evaluation. Two main tasks were performed in the present study. First, the genotoxicity data from comet assay for 16 noncoated metal oxide nanomaterials with different core composition were collected. An evaluation criterion was applied to establish which of these individual lines of evidence were of sufficient quality and what weight could have been given to them in inferring genotoxic results. The collected data were surveyed on (1) minimum necessary characterization points for nanomaterials and (2) principals of correct comet assay testing for nanomaterials. Second, in this study the genotoxicity effect of metal oxide nanomaterials was investigated by quantitative nanostructure–activity relationship approach. A set of quantum-chemical descriptors was developed for all investigated metal oxide nanomaterials. A classification model based on decision tree was developed for the investigated dataset. Thus, three descriptors were identified as the most responsible factors for genotoxicity effect: heat of formation, molecular weight, and surface area of the oxide cluster based on the conductor-like screening model. Conclusively, the proposed genotoxicity assessment strategy is useful to prioritize the study of the nanomaterials for further risk assessment evaluations.

  • Modeling gold nanoparticle biodistribution after arterial infusion into perfused tissue: effects of surface coating, size and protein corona
    Nanotoxicology (IF 5.811) Pub Date : 2018-06-01
    Jim E. Riviere, Majid Jaberi-Douraki, James Lillich, Tahmineh Azizi, Hyun Joo, Kyoungju Choi, Ravi Thakkar, Nancy A. Monteiro-Riviere

    A detailed understanding of the factors governing nanomaterial biodistribution is needed to rationally design safe nanomedicines. This research details the pharmacokinetics of gold nanoparticle (AuNP) biodistribution after arterial infusion of 40 or 80 nm AuNP (1 μg/ml) into the isolated perfused porcine skin flap (IPPSF). AuNP had surface coatings consisting of neutral polyethylene glycol (PEG), anionic lipoic acid (LA), or cationic branched polyethylenimine (BPEI). Effect of a porcine plasma corona (PPC) on 40 nm BPEI and PEG-AuNP were assessed in the IPPSF. Au concentrations were determined by ICP/MS and arterial to venous concentration-time profiles were analyzed over 8 hr (4 hr infusion, 4 hr washout) using a two-compartment pharmacokinetic model. IPPSF viability and vascular function were assessed by change in glucose utilization, vascular resistance, or weight gain after perfusion. All AuNP demonstrated some degree of AuNP arterial extraction and skin flap retention, as well as enhanced kinetic parameters of tissue uptake; with BPEI-AuNP consistently having the greatest biodistribution even with a PPC. Toxicological effects were not detected. Transmission electron microscopy confirmed intracellular uptake of AuNP. These studies paralleled previous in vitro cell culture studies using the same AuNP in human endothelial and renal proximal tubule cells, hepatocytes, keratinocytes, showing BPEI-AuNP having the greatest uptake, although the presence of a PPC did not reduce IPPSF biodistribution as in the cell culture studies. These findings clearly indicate arterial to the venous extraction of AuNP after infusion with the magnitude of extraction being greatest with the BPEI surface coating and provide data and model structure necessary to construct the whole body physiologically based pharmacokinetic models capable of utilizing available in vitro data.

  • The Enchytraeus crypticus stress metabolome – CuO NM case study
    Nanotoxicology (IF 5.811) Pub Date : 2018-06-22
    Vera L. Maria, David Licha, Christina Ranninger, Janeck J. Scott-Fordsmand, Christian G. Huber, Mónica J. B. Amorim

    The stress metabolome provides a thorough insight into the signals and hence mechanisms of response of organisms. This is an excellent tool to advance the understanding of interactions, especially for substances like nanomaterials (NMs), for which there is an urgent need for alternative methods for hazard assessment. The metabolome of Enchytraeus crypticus was studied for the first time. The case study, CuO NM (and CuCl2) covered exposure along a time frame [0–7–14 days (d)] and two reproduction effect concentrations (EC10 and EC50). High-performance liquid chromatography-mass spectrometry based method (HPLC-MS) was used, with reversed phase (RP) separation and mass spectrometric detection in positive and negative modes. Metabolite profiling of Cu materials yielded 155 and 382 metabolite features in positive and negative modes, respectively, showing an expression related with time, material, and ECx. The number of differentially expressed metabolites (DEMs) decreased with exposure time (14 d) for CuO NM, whereas for CuCl2 EC50 it increased. Overall, almost all DEMs are down-regulated for CuO NM and up-regulated for CuCl2 (both modes). Early effects were mainly related to amino acids and later to lysophospholipids (down-regulation). Furthermore, the underlying mechanisms of CuO NM toxicity (e.g. neurotransmission, nucleic acids generation, cellular energy, and immune defense) differ from CuCl2, where later metabolomic responses are mostly linked to the metabolism of lipids and fewer to amino acids. This study reports a large scale metabolome profiling for E. crypticus and identifies potential markers of Cu materials, which can help to align intelligent testing strategies and safer-by-design materials.

  • Co-exposure to silver nanoparticles and cadmium induce metabolic adaptation in HepG2 cells
    Nanotoxicology (IF 5.811) Pub Date : 2018-07-11
    Renata Rank Miranda, Vladimir Gorshkov, Barbara Korzeniowska, Stefan J. Kempf, Francisco Filipak Neto, Frank Kjeldsen

    Although multiple studies have reported the toxicological effects and underlying mechanisms of toxicity of silver nanoparticles (AgNP) in a variety of organisms, the interactions of AgNP with environmental contaminants such as cadmium are poorly understood. We used biochemical assays and mass spectrometry-based proteomics to assess the cellular and molecular effects induced by a co-exposure of HepG2 cells to AgNP and cadmium. Cell viability and energy homeostasis were slightly affected after a 4-h exposure to AgNP, cadmium, or a combination of the two; these endpoints were substantially altered after a 24-h co-exposure to AgNP and cadmium, while exposure to one of the two contaminants led only to minor changes. Proteomics analysis followed the same trend: while a 4-h exposure induced minor protein deregulation, a 24-h exposure to a combination of AgNP and cadmium deregulated 43% of the proteome. The toxicity induced by a combined exposure to AgNP and cadmium involved (1) inactivation of Nrf2, resulting in downregulation of antioxidant defense and proteasome-related proteins, (2) metabolic adaptation and ADP/ATP imbalance, and (3) increased protein synthesis possibly to reestablish homeostasis. The adaptation strategy was not sufficient to restore ADP/ATP homeostasis and to avoid cell death.

  • Development of a systematic method to assess similarity between nanomaterials for human hazard evaluation purposes – lessons learnt
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-06
    Margriet VDZ Park, Julia Catalán, Natalia Ferraz, Joan Cabellos, Ralph Vanhauten, Socorro Vázquez-Campos, Gemma Janer

    Within the EU FP-7 GUIDEnano project, a methodology was developed to systematically quantify the similarity between a nanomaterial (NM) that has been tested in toxicity studies and the NM for which risk needs to be evaluated, for the purpose of extrapolating toxicity data between the two materials. The methodology is a first attempt to use current knowledge on NM property–hazard relationships to develop a series of pragmatic and systematic rules for assessing NM similarity. Moreover, the methodology takes into account the practical feasibility, in that it is based on generally available NM characterization information. In addition to presenting this methodology, the lessons learnt and the challenges faced during its development are reported here. We conclude that there is a large gap between the information that is ideally needed and its application to real cases. The current database on property–hazard relationships is still very limited, which hinders the agreement on the key NM properties constituting the basis of the similarity assessment and the development of associated science-based and unequivocal rules. Currently, one of the most challenging NM properties to systematically assess in terms of similarity between two NMs is surface coating and functionalization, which lacks standardized parameters for description and characterization methodology. Standardization of characterization methods that lead to quantitative, unambiguous, and measurable parameters describing NM properties are necessary in order to build a sufficiently robust property–hazard database that allows for evidence-based refinement of our methodology, or any other attempt to systematically assess the similarity of NMs.

  • Quantitation of cell-associated carbon nanotubes: selective binding and accumulation of carboxylated carbon nanotubes by macrophages
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-26
    Ruhung Wang, Michael Lee, Karina Kinghorn, Tyler Hughes, Ishwar Chuckaree, Rishabh Lohray, Erik Chow, Paul Pantano, Rockford Draper

    To understand the influence of carboxylation on the interaction of carbon nanotubes with cells, the amount of pristine multi-walled carbon nanotubes (P-MWNTs) or carboxylated multi-walled carbon nanotubes (C-MWNTs) coated with Pluronic® F-108 that were accumulated by macrophages was measured by quantifying CNTs extracted from cells. Mouse RAW 264.7 macrophages and differentiated human THP-1 (dTHP-1) macrophages accumulated 80–100 times more C-MWNTs than P-MWNTs during a 24-h exposure at 37 °C. The accumulation of C-MWNTs by RAW 264.7 cells was not lethal; however, phagocytosis was impaired as subsequent uptake of polystyrene beads was reduced after a 20-h exposure to C-MWNTs. The selective accumulation of C-MWNTs suggested that there might be receptors on macrophages that bind C-MWNTs. The binding of C-MWNTs to macrophages was measured as a function of concentration at 4 °C in the absence of serum to minimize the potential interference by serum proteins or temperature-dependent uptake processes. The result was that the cells bound 8.7 times more C-MWNTs than P-MWNTs, consistent with the selective accumulation of C-MWNTs at 37 °C. In addition, serum strongly antagonized the binding of C-MWTS to macrophages, suggesting that serum contained inhibitors of binding. Moreover, inhibitors of class A scavenger receptor (SR-As) reduced the binding of C-MWNTs by about 50%, suggesting that SR-As contribute to the binding and endocytosis of C-MWNTs in macrophages but that other receptors may also be involved. Altogether, the evidence supports the hypothesis that macrophages contain binding sites selective for C-MWNTs that facilitate the high accumulation of C-MWNTs compared to P-MWNTs.

  • Acute effects of multi-walled carbon nanotubes on primary bronchial epithelial cells from COPD patients
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-26
    Seraina Beyeler, Savvina Chortarea, Barbara Rothen-Rutishauser, Alke Petri-Fink, Peter Wick, Stefan A. Tschanz, Christophe von Garnier, Fabian Blank

    The risks of occupational exposure during handling of multi-walled carbon nanotubes (MWCNTs) have received limited attention to date, in particular for potentially susceptible individuals with highly prevalent chronic obstructive pulmonary disease (COPD). In this in vitro study, we simulated acute inhalation of MWCNTs employing an air–liquid interface cell exposure (ALICE) system: primary human bronchial epithelial cells from COPD patients and healthy donors (controls), cultured at the air–liquid interface (ALI) were exposed to MWCNTs. To study acute health effects on the respiratory epithelium, two different concentrations (0.16; 0.34 µg/cm2) of MWCNTs were aerosolized onto cell cultures followed by analysis after 24 h. Following MWCNT exposure, epithelial integrity and differentiation remained intact. Electron microscopy analyses identified MWCNTs both extra- and intracellular within vesicles of mucus producing cells. In both COPD and healthy control cultures, MWCNTs neither caused increased release of lactate dehydrogenase (LDH), nor alterations in inflammatory responses, as measured by RNA expression and protein secretion of the cytokines IL-6, IL-8, CXCL10, IL-1β and TGF-β and oxidative stress markers HMOX-1 and SOD-2. No short-term alteration of epithelial cell function, as determined by ciliary beating frequency (CBF), occurred in any of the conditions tested. In conclusion, the present study provided a reliable and realistic in vitro acute-exposure model of the respiratory tract, responsive to positive controls such as Dörentruper Quartz (DQ12) and asbestos. Acute exposure to MWCNTs did not affect epithelial integrity, nor induce increased cell death, apoptosis or inflammatory changes.

  • High inflammogenic potential of rare earth oxide nanoparticles: the New Hazardous Entity
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-30
    Youngju Han, Dong-Keon Lee, Sung-Hyun Kim, Seonghan Lee, Soyeon Jeon, Wan-Seob Cho

    Due to the exponential increase in the development and utilization of rare earth oxide nanoparticles (REO NPs) in various fields, the possibility of exposure in humans by inhalation has increased. However, there are little information about hazards of REO NPs and its mechanisms of toxicity. In this study, we evaluated the acute pulmonary inflammation using 10 REO NPs (Dy2O3, Er2O3, Eu2O3, Gd2O3 La2O3, Nd2O3, Pr6O11, Sm2O3, Tb4O7, and Y2O3) and four well-known toxic particles (CuO, NiO, ZnO, and DQ12). Minimum three doses per NP were instilled into the lungs of female Wistar rats at surface area dose metric and lung inflammation was evaluated at 24 h post-instillation by bronchoalveolar lavage fluid (BALF) analysis and histopathological observation. All types of REO NPs showed common pathological changes including mild to moderate infiltration of neutrophils and activated macrophages in the alveoli, peribronchial, and perivascular region. The inflammogenic potential evaluated by the number of granulocytes divided by the treated surface area dose showed all types of REO NPs has much higher inflammogenic potential than DQ12, ZnO, and NiO NPs. The correlation plot between the number of granulocytes and the potential for reactive oxygen species (ROS) generation showed a good correlation with exception of Pr6O11. The higher inflammogenic potential of REO NPs than that of well-known highly toxic particles imply that REO NPs need special attention for inhalation exposure and more studies are needed. In addition, the potential of ROS generation is one of the key factors producing lung inflammation by REO NPs.

  • Nanofibrillated cellulose causes acute pulmonary inflammation that subsides within a month
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-30
    Marit Ilves, Sara Vilske, Kukka Aimonen, Hanna K. Lindberg, Saila Pesonen, Irene Wedin, Markus Nuopponen, Esa Vanhala, Casper Højgaard, Jakob R. Winther, Martin Willemoës, Ulla Vogel, Henrik Wolff, Hannu Norppa, Kai Savolainen, Harri Alenius

    Nanofibrillated cellulose (NFC) is a renewable nanomaterial that has beneficial uses in various applications such as packaging materials and paper. Like carbon nanotubes (CNT), NFCs have high aspect ratio and favorable mechanical properties. The aspect ratio also rises a concern whether NFC could pose a health risk and induce pathologies, similar to those triggered by multi-walled CNT. In this study, we explored the immunomodulatory properties of four NFCs in vitro and in vivo, and compared the results with data on bulk-sized cellulose fibrils and rigid multi-walled CNT (rCNT). Two of the NFCs were non-functionalized and two were carboxymethylated or carboxylated. We investigated the production of pro-inflammatory cytokines in differentiated THP-1 cells, and studied the pulmonary effects and biopersistence of the materials in mice. Our results demonstrate that one of the non-functionalized NFCs tested reduced cell viability and triggered pro-inflammatory reactions in vitro. In contrast, all cellulose materials induced innate immunity response in vivo 24 h after oropharyngeal aspiration, and the non-functionalized NFCs additionally caused features of Th2-type inflammation. Modest immune reactions were also seen after 28 days, however, the effects were markedly attenuated as compared with the ones after 24 h. Cellulose materials were not cleared within 1 month, as demonstrated by their presence in the exposed lungs. All effects of NFC were modest as compared with those induced by rCNT. NFC-induced responses were similar or exceeded those triggered by bulk-sized cellulose. These data provide new information about the biodurability and pulmonary effects of different NFCs; this knowledge can be useful in the risk assessment of cellulose materials.

  • Quantitative human health risk assessment along the lifecycle of nano-scale copper-based wood preservatives
    Nanotoxicology (IF 5.811) Pub Date : 2018-06-12
    Danail Hristozov, Lisa Pizzol, Gianpietro Basei, Alex Zabeo, Aiga Mackevica, Steffen Foss Hansen, Ilse Gosens, Flemming R. Cassee, Wim de Jong, Antti Joonas Koivisto, Nicole Neubauer, Araceli Sanchez Jimenez, Elena Semenzin, Vrishali Subramanian, Wouter Fransman, Keld Alstrup Jensen, Wendel Wohlleben, Vicki Stone, Antonio Marcomini

    The use of nano-scale copper oxide (CuO) and basic copper carbonate (Cu2(OH)2CO3) in both ionic and micronized wood preservatives has raised concerns about the potential of these substances to cause adverse humans health effects. To address these concerns, we performed quantitative (probabilistic) human health risk assessment (HHRA) along the lifecycles of these formulations used in antibacterial and antifungal wood coatings and impregnations by means of the EU FP7 SUN project’s Decision Support System (SUNDS, www.sunds.gd). The results from the risk analysis revealed inhalation risks from CuO in exposure scenarios involving workers handling dry powders and performing sanding operations as well as potential ingestion risks for children exposed to nano Cu2(OH)2CO3 in a scenario involving hand-to-mouth transfer of the substance released from impregnated wood. There are, however, substantial uncertainties in these results, so some of the identified risks may stem from the safety margin of extrapolation to fill data gaps and might be resolved by additional testing. Our stochastic approach successfully communicated the contribution of different sources of uncertainty in the risk assessment. The main source of uncertainty was the extrapolation from short to long-term exposure, which was necessary due to the lack of (sub)chronic in vivo studies with CuO and Cu2(OH)2CO3. Considerable uncertainties also stemmed from the use of default inter- and intra-species extrapolation factors.

  • Multi-level toxicity assessment of engineered cellulose nanofibrils in Daphnia magna
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-06
    Martin Ogonowski, Ulrica Edlund, Elena Gorokhova, Margareta Linde, Karin Ek, Birgitta Liewenborg, Oda Könnecke, Julien R. G. Navarro, Magnus Breitholtz

    Cellulose nanofibril (CNF)-based materials are increasingly used in industrial and commercial applications. However, the impacts of CNF on aquatic life are poorly understood, and there are concerns regarding their potential toxicity. Using a combination of standard ecotoxicological tests and feeding experiments, we assessed the effects of CNF exposure (0.206–20.6 mg/L) on the feeding (food uptake and gut residence time) and life-history traits (growth and reproduction) in the cladoceran Daphnia magna. No mortality was observed in a 48 h acute exposure at 2060 mg/L. Moreover, a 21-day exposure at low food and moderate CNF levels induced a stimulatory effect on growth, likely driven by increased filtration efficiency, and, possibly, partial assimilation of the CNF by the animals. However, at low food levels and the highest CNF concentrations, growth and reproduction were negatively affected. These responses were linked to caloric restriction caused by dilution of the food source, but not an obstruction of the alimentary canal. Finally, no apparent translocation of CNF past the alimentary canal was detected. We conclude that CNF displays a low toxic potential to filter-feeding organisms and the expected environmental risks are low.

  • Inflammation in the pleural cavity following injection of multi-walled carbon nanotubes is dependent on their characteristics and the presence of IL-1 genes
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-09
    Yke Jildouw Arnoldussen, Vidar Skaug, Mona Aleksandersen, Erik Ropstad, Kristine Haugen Anmarkrud, Elin Einarsdottir, Fang Chin-Lin, Cesilie Granum Bjørklund, Mayes Kasem, Einar Eilertsen, Ron N. Apte, Shanbeh Zienolddiny

    Upon inhalation, multi-walled carbon nanotubes (MWCNTs) may reach the subpleura and pleural spaces, and induce pleural inflammation and/or mesothelioma in humans. However, the mechanisms of MWCNT-induced pathology after direct intrapleural injections are still only partly elucidated. In particular, a role of the proinflammatory interleukin-1 (IL-1) cytokines in pleural inflammation has so far not been published. We examined the MWCNT-induced pleural inflammation, gene expression abnormalities, and the modifying role of IL-1α and β cytokines following intrapleural injection of two types of MWCNTs (CNT-1 and CNT-2) compared with crocidolite asbestos in IL-1 wild-type (WT) and IL-1α/β KO (IL1-KO) mice. Histopathological examination of the pleura 28 days post-exposure revealed mesothelial cell hyperplasia, leukocyte infiltration, and fibrosis occurring in the CNT-1 (Mitsui-7)-exposed group. The pleura of these mice also showed the greatest changes in mRNA and miRNA expression levels, closely followed by CNT-2. In addition, the CNT-1-exposed group also presented the greatest infiltrations of leukocytes and proliferation of fibrous tissue. WT mice were more prone to development of sustained inflammation and fibrosis than IL1-KO mice. Prominent differences in genetic and epigenetic changes were also observed between the two genotypes. In conclusion, the fibrotic response to MWCNTs in the pleura depends on the particles’ physico-chemical properties and on the presence or absence of the IL-1 genes. Furthermore, we found that CNT-1 was the most potent inducer of inflammatory responses, followed by CNT-2 and crocidolite asbestos.

  • The small airway epithelium as a target for the adverse pulmonary effects of silver nanoparticle inhalation
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-11
    Chang Guo, Alison Buckley, Tim Marczylo, Joanna Seiffert, Isabella Römer, James Warren, Alan Hodgson, Kian Fan Chung, Timothy W. Gant, Rachel Smith, Martin O. Leonard

    Experimental modeling to identify specific inhalation hazards for nanomaterials has in the main focused on in vivo approaches. However, these models suffer from uncertainties surrounding species-specific differences and cellular targets for biologic response. In terms of pulmonary exposure, approaches which combine ‘inhalation-like’ nanoparticulate aerosol deposition with relevant human cell and tissue air–liquid interface cultures are considered an important complement to in vivo work. In this study, we utilized such a model system to build on previous results from in vivo exposures, which highlighted the small airway epithelium as a target for silver nanoparticle (AgNP) deposition. RNA-SEQ was used to characterize alterations in mRNA and miRNA within the lung. Organotypic-reconstituted 3D human primary small airway epithelial cell cultures (SmallAir) were exposed to the same spark-generated AgNP and at the same dose used in vivo, in an aerosol-exposure air–liquid interface (AE-ALI) system. Adverse effects were characterized using lactate, LDH release and alterations in mRNA and miRNA. Modest toxicological effects were paralleled by significant regulation in gene expression, reflective mainly of specific inflammatory events. Importantly, there was a level of concordance between gene expression changes observed in vitro and in vivo. We also observed a significant correlation between AgNP and mass equivalent silver ion (Ag+) induced transcriptional changes in SmallAir cultures. In addition to key mechanistic information relevant for our understanding of the potential health risks associated with AgNP inhalation exposure, this work further highlights the small airway epithelium as an important target for adverse effects.

  • Elucidating differential nano-bio interactions of multi-walled andsingle-walled carbon nanotubes using subcellular proteomics
    Nanotoxicology (IF 5.811) Pub Date : 2018-04-24
    Joseph D. T. Ndika, Jukka Sund, Harri Alenius, Anne Puustinen

    Understanding the relationship between adverse exposure events and specific material properties will facilitate predictive classification of carbon nanotubes (CNTs) according to their mechanisms of action, and a safe-by-design approach for the next generation of CNTs. Mass-spectrometry-based proteomics is a reliable tool to uncover the molecular dynamics of hazardous exposures, yet challenges persist with regards to its limited dynamic range when sampling whole organisms, tissues or cell lysates. Here, the simplicity of the sub-cellular proteome was harnessed to unravel distinctive adverse exposure outcomes at the molecular level, between two CNT subtypes. A549, MRC9 and human macrophage cells, were exposed for 24h to non-cytotoxic doses of single-walled or multi-walled CNTs (swCNTs or mwCNTs). Label-free proteomics on enriched cytoplasmic fractions was complemented with analyses of reactive oxygen species (ROS) production and mitochondrial integrity. The extent/number of modulated proteoforms indicated the single-walled variant was more bioactive. Greater enrichment of pathways corresponding to oxido-reductive activity was consistent with greater intracellular ROS induction and mitochondrial dysfunction capacities of swCNTs. Other compromised cellular functions, as revealed by pathway analysis were; ribosome, spliceosome and DNA repair. Highly upregulated proteins (fold change in abundance >6) such as, APOC3, RBP4 and INS are also highlighted as potential markers of hazardous CNT exposure. We conclude that, changes in cytosolic proteome abundance resulting from nano-bio interactions, elucidate adverse response pathways and their distinctive molecular components. Our results indicate that CNT-protein interactions might have a thus far unappreciated significance for protein trafficking, and this warrants further investigation.

  • Pulmonary hypofunction due to calcium carbonate nanomaterial exposure in occupational workers: a cross-sectional study
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-06
    Guoliang Li, Lihong Liang, Jingchao Yang, Lihai Zeng, Zhiwei Xie, Yizhou Zhong, Xiaolin Ruan, Ming Dong, Zhanhong Yang, Guanchao Lai, Weixin Huang, Aichu Yang, Jiabing Chen, Banghua Wu, Huaming Xu, Dezhi Meng, Shijie Hu, Lihua Xia, Xingfen Yang, Laiyu Li, Sahoko Ichihara, Gaku Ichihara, Hanlin Huang, Zhenlie Huang

    Calcium carbonate nanomaterials (nano-CaCO3) are widely used in both manufacturing and consumer products, but their potential health hazards remain unclear. The objective of this study was to survey workplace exposure levels and health effects of workers exposed to nano-CaCO3. Personal and area sampling, as well as real-time and dust monitoring, were performed to characterize mass exposure, particle size distribution, and particle number exposure. A total of 56 workers (28 exposed workers and 28 unexposed controls) were studied in a cross-sectional study. They completed physical examinations, spirometry, and digital radiography. The results showed that the gravimetric nano-CaCO3 concentration was 5.264 ± 6.987 mg/m3 (0.037–22.192 mg/m3) at the workplace, and 3.577 ± 2.065 mg/m3 (2.042–8.161 mg/m3) in the breathing zone of the exposed workers. The particle number concentrations ranged from 8193 to 39 621 particles/cm3 with a size range of 30–150 nm. The process of packing had the highest gravimetric and particle number concentrations. The particle number concentration positively correlated with gravimetric concentrations of nano-CaCO3. The levels of hemoglobin, creatine phosphokinase (CK), lactate dehydrogenase, and high-density lipoprotein cholesterol (HDL-C) in the nano-CaCO3 exposure group increased significantly, but the white blood cell count (WBC), Complement 3 (C3), total protein (TP), uric acid, and creatinine (CREA) all decreased significantly. The prevalence rate of pulmonary hypofunction was significantly higher (p = 0.037), and the levels of vital capacity (VC), forced vital capacity (FVC), forced expiratory volume in one second (FEV1), FEV1/FVC, peak expiratory flow and forced expiratory flow 25% (FEF 25%), FEF 25–75% were negatively correlated with gravimetric concentrations of nano-CaCO3 (p < 0.05). Logistic analysis showed that nano-CaCO3 exposure level was associated with pulmonary hypofunction (p = 0.005). Meanwhile, a dose-effect relationship was found between the accumulated gravimetric concentrations of nano-CaCO3 and the prevalence rate of pulmonary hypofunction (p = 0.048). In conclusion, long-term and high-level nano-CaCO3 exposure can induce pulmonary hypofunction in workers. Thus, lung function examination is suggested for occupational populations with nano-CaCO3 exposure. Furthermore, future health protection efforts should focus on senior workers with accumulation effects of nano-CaCO3 exposure.

  • The protective role of autophagy in nephrotoxicity induced by bismuth nanoparticles through AMPK/mTOR pathway
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-06
    Yongming Liu, Huan Yu, Xihui Zhang, Yong Wang, Zhentao Song, Jian Zhao, Haibin Shi, Ruibin Li, Yangyun Wang, Leshuai W. Zhang

    Bismuth is widely used in metallurgy, cosmetic industry, and medical diagnosis and recently, bismuth nanoparticles (NPs) (BiNP) have been made and proved to be excellent CT imaging agents. Previously, we have synthesized bovine serum albumin based BiNP for imaging purpose but we found a temporary kidney injury by BiNP. Due to the reported adverse events of bismuth on human health, we extended our studies on the mechanisms for BiNP induced nephrotoxicity. Blood biochemical analysis indicated the increase in creatinine (CREA) and blood urea nitrogen (BUN), and intraluminal cast formation with cell apoptosis/necrosis was evident in proximal convoluted tubules (PCTs) of mice. BiNP induced acute kidney injury (AKI) was associated with an increase in LC3II, while the autophagic flux indicator p62 remained unchanged. Chloroquine and rapamycin were used to evaluate the role of autophagy in AKI caused by BiNP. Results showed that BiNP induced AKI was further attenuated by rapamycin, while AKI became severe when chloroquine was applied. In vitro studies further proved BiNP induced autophagy in human embryonic kidney cells 293, presented as autophagic vacuole (AV) formation along with increased levels of autophagy-related proteins including LC3II, Beclin1, and Atg12. Specifically, reactive oxygen species (ROS) generated by BiNP could be the major inducer of autophagy, because ROS blockage attenuated autophagy. Autophagy induced by BiNP was primarily regulated by AMPK/mTOR signal pathway and partially regulated by Akt/mTOR. Our study provides fundamental theory to better understand bismuth induced nephrotoxicity for better clinical application of bismuth related compounds.

  • Mechanistic insight into reactivity and (geno)toxicity of well-characterized nanoparticles of cobalt metal and oxides
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-23
    Francesca Cappellini, Yolanda Hedberg, Sarah McCarrick, Jonas Hedberg, Remco Derr, Giel Hendriks, Inger Odnevall Wallinder, Hanna L. Karlsson

    An increasing use of cobalt (Co)-based nanoparticles (NPs) in different applications and exposures at occupational settings triggers the need for toxicity assessment. Improved understanding regarding the physiochemical characteristics of Co metal NPs and different oxides in combination with assessment of toxicity and mechanisms may facilitate decisions for grouping during risk assessment. The aim of this study was to gain mechanistic insights in the correlation between NP reactivity and toxicity of three different Co-based NPs (Co, CoO, and Co3O4) by using various tools for characterization, traditional toxicity assays, as well as six reporter cell lines (ToxTracker) for rapid detection of signaling pathways of relevance for carcinogenicity. The results showed cellular uptake of all NPs in lung cells and induction of DNA strand breaks and oxidative damage (comet assay) by Co and CoO NPs. In-depth studies on the ROS generation showed high reactivity of Co, lower for CoO, and no reactivity of Co3O4 NPs. The reactivity depended on the corrosion and transformation/dissolution properties of the particles and the media highlighting the role of the surface oxide and metal speciation as also confirmed by in silico modeling. By using ToxTracker, Co NPs were shown to be highly cytotoxic and induced reporters related to oxidative stress (Nrf2 signaling) and DNA strand breaks. Similar effects were observed for CoO NPs but at higher concentrations, whereas the Co3O4 NPs were inactive at all concentrations tested. In conclusion, our study suggests that Co and CoO NPs, but not Co3O4, may be grouped together for risk assessment.

  • Metabolic fate and subchronic biological effects of core–shell structured Fe3O4@SiO2-NH2 nanoparticles
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-10
    Yueli Chen, Jinquan Li, Zhongxue Yuan, Jianghua Feng, Zhong Chen

    Core–shell structured Fe3O4@SiO2-NH2 nanoparticles (Fe@Si-NPs) demonstrated outstanding potentials in drug targeting and delivery and medical imaging. However, they have limited clinical applications due to unknown chronic bio-effects and potential bio-related risks. In this study, the subchronic biological effects and metabolic fate of 20 nm Fe@Si-NPs in Sprague–Dawley rats in 12 weeks were investigated by the biochemical assay and NMR-based metabonomic analysis using an intravenous model. Biofluids (plasma and urine) analysis provided the transportation, absorption, and excretion information of Fe@Si-NPs. Urine metabonome displayed a metabolic recovery while self-regulation of plasma metabonome leaded to the parallel metabolic trends between dosed and control groups in 12 weeks. And biological tissues (spleen, liver, kidney, and lung) analysis indicated liver and spleen are the targeted-organs of Fe@Si-NPs. The obvious metabolic variations responding to the biodistribution were induced by Fe@Si-NPs although no visible toxic effects were observed in these tissues. Besides the common energy metabolism response to the xenobiotics, Fe@Si-NPs also disturbed the metabolic pathways in glycerophospholipid and sphingolipid metabolism, metabolisms of purine, pyrimidine, and nicotinate. Our results provide preliminary validation for the potential use of Fe@Si-NPs in clinical medicine and give identifiable ground for the dose selection and bio-nanoagent optimization.

  • Copper nanoparticles induce early fibrotic changes in the liver via TGF-β/Smad signaling and cause immunosuppressive effects in rats
    Nanotoxicology (IF 5.811) Pub Date : 2018-05-30
    In-Chul Lee, Je-Won Ko, Sung-Hyeuk Park, Na-Rae Shin, In-Sik Shin, Changjong Moon, Sung-Ho Kim, Won-Kee Yun, Hyoung-Chin Kim, Jong-Choon Kim

    Copper nanoparticles (Cu NPs) have various uses, including as additives in polymers/plastics, lubricants for metallic coating, and biomedical applications. We investigated the role of transforming growth factor (TGF)-β1 signaling in hepatic damage caused by Cu NPs and explored the effects of a 28-day repeated oral administration to Cu NPs on the immune response. The exposure to Cu NPs caused a dose-dependent increase in Cu levels in the liver and spleen. Cu NPs caused hepatic damage and markedly increased oxidative stress in liver tissues. Cu NPs induced activation of TGF-β1/Smad signaling by induction of vascular endothelial growth factor and matrix metalloproteinase-9. Exposure to Cu NPs also induced activation of Smad-independent pathways, phosphorylation of mitogen-activated protein kinases (MAPKs) and Akt/FoxO3. Consistent with the activation of TGF-β1/Smad-dependent and -independent pathways, Cu NPs markedly increased the deposition and induction of extracellular matrix components, α-smooth muscle actin, and collagens in liver tissues. In addition, repeated exposure to Cu NPs suppressed the proliferation of mitogenically stimulated T- or B-lymphocytes and decreased CD3+ (particularly, CD3+CD4+CD8−) and CD45+ population, followed by decreased levels of immunoglobulins and Th1/Th2 type cytokines. Collectively, Cu NPs caused hepatic damage and induced pro-fibrotic changes, which were closely related to the activation of oxidative stress-mediated TGF-β1/Smad-dependent and -independent pathways (MAPKs and Akt/FoxO3). We confirmed the immunosuppressive effect of Cu NPs via the inhibition of mitogen-stimulated spleen-derived lymphocyte proliferation and suppression of B- or T-lymphocyte-mediated immune responses.

  • Comparative study of dissolved and nanoparticulate Ag effects on the life cycle of an estuarine meiobenthic copepod, Amphiascus tenuiremis
    Nanotoxicology (IF 5.811) Pub Date : 2018-03-19
    Mithun Sikder, Emily Eudy, G. Thomas Chandler, Mohammed Baalousha

    Many nanotoxicological studies have assessed the acute toxicity of nanoparticles (NPs) at high exposure concentrations. There is a gap in understanding NP chronic environmental effects at lower exposure concentrations. This study reports life-cycle chronic toxicity of sublethal exposures of polyvinylpyrrolidone-coated silver nanoparticles (PVP-AgNPs) relative to dissolved silver nitrate (AgNO3) for the estuarine meiobenthic copepod, Amphiascus tenuiremis, over a range of environmentally relevant concentrations, i.e., 20, 30, 45, and 75 µg-Ag L−1. A concentration-dependent increase in mortality of larval nauplii and juvenile copepodites was observed. In both treatment types, significantly higher mortality was observed at 45 and 75 µg-Ag L−1 than in controls. In AgNO3 exposures, fecundity declined sharply (1.8–7 fold) from 30 to 75 µg Ag L−1. In contrast, fecundity was not affected by PVP-AgNPs exposures. A Leslie matrix population-growth model predicted sharply 60–86% of decline in overall population sizes and individual life-stage numbers from 30–75 µg-Ag L−1 as dissolved AgNO3. In contrast, no population growth suppressions were predicted for any PVP-AgNPs exposures. Slower release of dissolved Ag from PVP-AgNPs and/or reduced Ag uptake in the nanoform may explain these sharp contrasts in copepod response.

  • Titanium dioxide nanoparticle exposure alters metabolic homeostasis in a cell culture model of the intestinal epithelium and Drosophila melanogaster
    Nanotoxicology (IF 5.811) Pub Date : 2018-03-30
    Jonathan W. Richter, Gabriella M. Shull, John H. Fountain, Zhongyuan Guo, Laura P. Musselman, Anthony C. Fiumera, Gretchen J. Mahler

    Nanosized titanium dioxide (TiO2) is a common additive in food and cosmetic products. The goal of this study was to investigate if TiO2 nanoparticles affect intestinal epithelial tissues, normal intestinal function, or metabolic homeostasis using in vitro and in vivo methods. An in vitro model of intestinal epithelial tissue was created by seeding co-cultures of Caco-2 and HT29-MTX cells on a Transwell permeable support. These experiments were repeated with monolayers that had been cultured with the beneficial commensal bacteria Lactobacillus rhamnosus GG (L. rhamnosus). Glucose uptake and transport in the presence of TiO2 nanoparticles was assessed using fluorescent glucose analog 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (2-NBDG). When the cell monolayers were exposed to physiologically relevant doses of TiO2, a statistically significant reduction in glucose transport was observed. These differences in glucose absorption were eliminated in the presence of beneficial bacteria. The decrease in glucose absorption was caused by damage to intestinal microvilli, which decreased the surface area available for absorption. Damage to microvilli was ameliorated in the presence of L. rhamnosus. Complimentary studies in Drosophila melanogaster showed that TiO2 ingestion resulted in decreased body size and glucose content. The results suggest that TiO2 nanoparticles alter glucose transport across the intestinal epithelium, and that TiO2 nanoparticle ingestion may have physiological consequences.

  • Improving colloidal stability of silica nanoparticles when stored in responsive gel: application and toxicity study
    Nanotoxicology (IF 5.811) Pub Date : 2018-04-02
    Giorgia Giovannini, Paul Warncke, Dagmar Fischer, Ondrej Stranik, Andrew J. Hall, Vladimir Gubala

    When silica nanoparticles (SiNP) are stored in aqueous solution, even for few hours, they have a tendency to form agglomerates and therefore adapt inhomogeneous structures. Here we present a very practical method to store SiNP in responsive hydrogel. We have confirmed that SiNP kept in the responsive hydrogel do not undergo through undesirable morphological changes and while in storage they maintain their excellent colloidal stability. The effect of SiNP hollowing (i.e. dissolution of the core of the particles that leaves empty cavity inside) was significantly inhibited in the hydrogel, which is a critical feature for any nano-medical applications (e.g. controlled drug release). To demonstrate the applicability of the hydrogel-storing concept within a biologically relevant context, in this work we have evaluated the toxicological effects of the responsive SiNP-gel formulation in a model in vitro (human cell line U87GM and hemocompatibility using red blood cells) and ex ovo (hen’s egg test) experiments. Particles stored in the gel as well as the pure gel did not affect the hemocompatibility (hemolysis and erythrocyte aggregation) up to a concentration of 100 µg/mL. Furthermore, systemic injections into the blood circulation of the chick area vasculosa confirmed the biocompatibility in a more complex biological environment. All evaluated toxicological values (hemorrhage, thrombosis, vascular lysis, and lethality) were comparable with the negative control, and no differences in toxicological response could be observed between the SiNP stored in hydrogel and the control nanoparticles stored in the solution.

  • Toxicity of mixtures of zinc oxide and graphene oxide nanoparticles to aquatic organisms of different trophic level: particles outperform dissolved ions
    Nanotoxicology (IF 5.811) Pub Date : 2018-04-14
    Nan Ye, Zhuang Wang, Se Wang, Willie J. G. M. Peijnenburg

    Concomitant releases of various engineered nanoparticles (NPs) into the environment have resulted in concerns regarding their combined toxicity to aquatic organisms. It is however, still elusive to distinguish the contribution to toxicity of components in NP mixtures. In the present study, we quantitatively evaluated the relative contribution of NPs in their particulate form (NP(particle)) and of dissolved ions released from NPs (NP(ion)) to the combined toxicity of binary mixtures of ZnO NPs and graphene oxide nanoplatelets (GO NPs) to three aquatic organisms of different trophic levels, including an alga species (Scenedesmus obliquus), a cladoceran species (Daphnia magna), and a freshwater fish larva (Danio rerio). Our results revealed that the effects of ZnO NPs and GO NPs were additive to S. obliquus and D. magna but antagonistic to D. rerio. The relative contribution to toxicity (RCT) of the mixture components to S. obliquus decreased in the order of RCTGO NP(particle) > RCTZnO NP(particle) > RCTZnO NP(ion), while the RCT of the mixture components to D. magna and D. rerio decreased in the order of RCTZnO NP(particle) > RCTGO NP(particle) > RCTZnO NP(ion). This finding also implies that the suspended particles rather than the dissolved Zn-ions dictated the combined toxicity of binary mixtures of ZnO NPs and GO NPs to the aquatic organisms of different trophic level. The alleviation of the contribution to toxicity of the ionic form of ZnO NPs was caused by the adsorption of the dissolved ions on GO NPs. Furthermore, the ZnO NP(particle) and GO NP(particle) displayed a different contribution to the observed mixture toxicity, dependent on the trophic level of the aquatic organisms tested. The difference of the contributions between the two particulate forms was mainly associated with differences in the intracellular accumulation of reactive oxygen species. Our findings highlight the important role of particles in the ecological impact of multi-nanomaterial systems.

  • LC-MS-based lipidomics to examine acute rat pulmonary responses after nano- and fine-sized ZnO particle inhalation exposure
    Nanotoxicology (IF 5.811) Pub Date : 2018-04-11
    Sheng-Han Lee, Chuan-Ho Tang, Wan-Yu Lin, Ke-Han Chen, Hao-Jan Liang, Tsun-Jen Cheng, Ching-Yu Lin

    Zinc oxide (ZnO) nano- and fine-sized particles are associated with respiratory toxicity in humans, but the underlying molecular mechanisms remain unclear. Our previous nuclear magnetic resonance-based metabolomic study demonstrated that changes in phosphorylcholine-containing lipids (PC-CLs) in the respiratory system were associated with ZnO particle-induced respiratory toxicity. However, the details of the lipid species associated with adverse effects and possible biomarker signatures have not been identified. Thus, a liquid chromatography-mass spectrometry (LC-MS)-based lipidomics platform was applied to examine the alterations of PC-CL species in the lungs of rats treated with a series of concentrations of nano-sized (35 nm) or fine-sized (250 nm) ZnO particles via inhalation. Principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and the Mann–Whitney U (MWU) test with false discovery rate (FDR) control were conducted to explore the perturbed lipid species and to discriminate a potential pulmonary biomarker signature after ZnO particle exposure. The PCA and PLS-DA models revealed that the fine-sized ZnO particle-treated groups and the high-concentration nano-sized group were separated from the control groups as well as from the low and moderate nano-sized groups. The results from the MWU test further suggested that after FDR adjustment, numerous PC-CL species were altered in the high-concentration and moderate-concentration fine-sized groups. Furthermore, our results suggested that lipids involved in anti-oxidation, membrane conformation, and cellular signal transduction were altered in response to ZnO-induced oxidative stress and inflammation. One lipid, PC(18:0/18:1), exhibited good performance (AUC > 0.8) of discriminative ability in distinguishing ZnO particle exposure from the control. These findings not only provide a foundation for the exploration of possible ZnO particle-mediated mechanisms but also suggest a lipid biomarker for ZnO particle exposure.

  • Probabilistic risk assessment of gold nanoparticles after intravenous administration by integrating in vitro and in vivo toxicity with physiologically based pharmacokinetic modeling
    Nanotoxicology (IF 5.811) Pub Date : 2018-04-14
    Yi-Hsien Cheng, Jim E. Riviere, Nancy A. Monteiro-Riviere, Zhoumeng Lin

    This study aimed to conduct an integrated and probabilistic risk assessment of gold nanoparticles (AuNPs) based on recently published in vitro and in vivo toxicity studies coupled to a physiologically based pharmacokinetic (PBPK) model. Dose–response relationships were characterized based on cell viability assays in various human cell types. A previously well-validated human PBPK model for AuNPs was applied to quantify internal concentrations in liver, kidney, skin, and venous plasma. By applying a Bayesian-based probabilistic risk assessment approach incorporating Monte Carlo simulation, probable human cell death fractions were characterized. Additionally, we implemented in vitro to in vivo and animal-to-human extrapolation approaches to independently estimate external exposure levels of AuNPs that cause minimal toxicity. Our results suggest that under the highest dosing level employed in existing animal studies (worst-case scenario), AuNPs coated with branched polyethylenimine (BPEI) would likely induce ∼90–100% cellular death, implying high cytotoxicity compared to <10% cell death induced by low-to-medium animal dosing levels, which are commonly used in animal studies. The estimated human equivalent doses associated with 5% cell death in liver and kidney were around 1 and 3 mg/kg, respectively. Based on points of departure reported in animal studies, the human equivalent dose estimates associated with gene expression changes and tissue cell apoptosis in liver were 0.005 and 0.5 mg/kg, respectively. Our analyzes provide insights into safety evaluation, risk prediction, and point of departure estimation of AuNP exposure for humans and illustrate an approach that could be applied to other NPs when sufficient data are available.

  • Inflammation–coagulation response and thrombotic effects induced by silica nanoparticles in zebrafish embryos
    Nanotoxicology (IF 5.811) Pub Date : 2018-04-14
    Junchao Duan, Shuang Liang, Yang Yu, Yang Li, Lijing Wang, Zehao Wu, Yueyue Chen, Mark R. Miller, Zhiwei Sun

    Nowadays, nanotechnology environmental health and safety (nanoEHS) is gaining attention. We previously found that silica nanoparticles (SiNPs) could induce vascular endothelial damage. However, the subsequent toxicologic response to SiNPs-induced endothelial damage was still largely unknown. In this study, we explored the inflammation–coagulation response and thrombotic effects of SiNPs in endothelial cells and zebrafish embryos. For in vitro study, swollen mitochondria and autophagosome were observed in ultrastructural analysis. The cytoskeleton organization was disrupted by SiNPs in vascular endothelial cells. The release of proinflammatory and procoagulant cytokines including IL-6, IL-8, MCP-1, PECAM-1, TF and vWF, were markedly elevated in a dose-dependent manner. For in vivo study, based on the NOAEL for dosimetry selection, and using two transgenic zebrafish, Tg(mpo:GFP) and Tg(fli-1:EGFP), SiNPs-induced neutrophil-mediated inflammation and impaired vascular endothelial cells. With the dosage higher than NOAEL, SiNPs significantly decreased blood flow and velocity, exhibiting a blood hypercoagulable state in zebrafish embryos. The thrombotic effect was assessed by o-dianisidine staining, showed that an increasing of erythrocyte aggregation occurred in SiNPs-treated zebrafish. Microarray analysis was used to screen the possible genes for inflammation–coagulation response to SiNPs in zebrafish, and the JAK1/TF signaling pathway was further verified by qRT-PCR and Western blot assays. For in-deepth study, il6st was knocked down with specific morpholinos. The whole-mount in situ hybridization and qRT-PCR analysis showed that the expression jak1 and f3b were attenuated in il6st knockdown groups. In summary, our data demonstrated that SiNPs could induce inflammation–coagulation response and thrombotic effects via JAK1/TF signaling pathway.

  • Silicon dioxide nanoparticle exposure affects small intestine function in an in vitro model
    Nanotoxicology (IF 5.811) Pub Date : 2018-04-18
    Zhongyuan Guo, Nicole J. Martucci, Yizhong Liu, Eusoo Yoo, Elad Tako, Gretchen J. Mahler

    The use of nanomaterials to enhance properties of food and improve delivery of orally administered drugs has become common, but the potential health effects of these ingested nanomaterials remain unknown. The goal of this study is to characterize the properties of silicon dioxide (SiO2) nanoparticles (NP) that are commonly used in food and food packaging, and to investigate the effects of physiologically realistic doses of SiO2 NP on gastrointestinal (GI) health and function. In this work, an in vitro model composed of Caco-2 and HT29-MTX co-cultures, which represent absorptive and goblet cells, was used. The model was exposed to well-characterized SiO2 NP for acute (4 h) and chronic (5 d) time periods. SiO2 NP exposure significantly affected iron (Fe), zinc (Zn), glucose, and lipid nutrient absorption. Brush border membrane intestinal alkaline phosphatase (IAP) activity was increased in response to nano-SiO2. The barrier function of the intestinal epithelium, as measured by transepithelial electrical resistance, was significantly decreased in response to chronic exposure. Gene expression and oxidative stress formation analysis showed NP altered the expression levels of nutrient transport proteins, generated reactive oxygen species, and initiated pro-inflammatory signaling. SiO2 NP exposure damaged the brush border membrane by decreasing the number of intestinal microvilli, which decreased the surface area available for nutrient absorption. SiO2 NP exposure at physiologically relevant doses ultimately caused adverse outcomes in an in vitro model.

  • Graphene onto medical grade titanium: an atom-thick multimodal coating that promotes osteoblast maturation and inhibits biofilm formation from distinct species
    Nanotoxicology (IF 5.811) Pub Date : 2018-02-06
    Nileshkumar Dubey, Kassapa Ellepola, Fanny E. D. Decroix, Julien L. P. Morin, AH Castro Neto, Chaminda J. Seneviratne, Vinicius Rosa

    The time needed for the osseointegration of titanium implants is deemed too long. Moreover, the bacterial colonization of their surfaces is a major cause of failure. Graphene can overcome these issues but its wet transfer onto substrates employs hazardous chemicals limiting the clinical applications. Alternatively, dry transfer technique has been developed, but the biological properties of this technique remain unexplored. Here, a dry transfer technique based on a hot-pressing method allowed to coat titanium substrates with high-quality graphene and coverage area >90% with a single transfer. The graphene-coated titanium is cytocompatible, did not induce cell membrane damage, induced human osteoblast maturation (gene and protein level), and increased the deposition of mineralized matrix compared to titanium alone. Moreover, graphene decreased the formation of biofilms from Streptococcus mutans, Enterococcus faecalis and even from whole saliva on titanium without killing the bacteria. These findings confirm that coating of titanium with graphene via a dry transfer technique is a promising strategy to improve osseointegration and prevent biofilm formation on implants and devices.

  • Impact of acute and subchronic inhalation exposure to PbO nanoparticles on mice
    Nanotoxicology (IF 5.811) Pub Date : 2018-02-15
    J. Lebedová, Z. Nováková, Z. Večeřa, M. Buchtová, J. Dumková, B. Dočekal, L. Bláhová, P. Mikuška, I. Míšek, A. Hampl, K. Hilscherová

    Lead nanoparticles (NPs) are released into air from metal processing, road transport or combustion processes. Inhalation exposure is therefore very likely to occur. However, even though the effects of bulk lead are well known, there is limited knowledge regarding impact of Pb NPs inhalation. This study focused on acute and subchronic exposures to lead oxide nanoparticles (PbO NPs). Mice were exposed to PbO NPs in whole body inhalation chambers for 4–72 h in acute experiment (4.05 × 106 PbO NPs/cm3), and for 1–11 weeks in subchronic experiment (3.83 × 105 particles/cm3 in lower and 1.93 × 106 particles/cm3 in higher exposure group). Presence of NPs was confirmed in all studied organs, including brain, which is very important considering lead neurotoxicity. Lead concentration gradually increased in all tissues depending on the exposure concentration and duration. The most burdened organs were lung and kidney, however liver and brain also showed significant increase of lead concentration during exposure. Histological analysis documented numerous morphological alterations and tissue damage, mainly in lung, but also in liver. Mild pathological changes were observed also in kidney and brain. Levels of glutathione (reduced and oxidized) were modulated mainly in lung in both, acute and subchronic exposures. Increase of lipid peroxidation was observed in kidney after acute exposure. This study characterized impacts of short to longer-term inhalation exposure, proved transport of PbO NPs to secondary organs, documented time and concentration dependent gradual increase of Pb concentration and histopathological damage in tissues.

  • The fate of silver nanoparticles in authentic human saliva
    Nanotoxicology (IF 5.811) Pub Date : 2018-02-16
    Kamonwad Ngamchuea, Christopher Batchelor-McAuley, Richard G. Compton

    The physicochemical properties of silver nanoparticles (AgNPs) in human whole saliva are investigated herein. In authentic saliva samples, AgNPs exhibit a great stability with over 70% of the nanomaterial remaining intact after a 24-h incubation in the presence of ∼0.3 mM dissolved oxygen. The small loss of AgNPs from the saliva sample has been demonstrated to be a result of two processes: agglomeration/aggregation (not involving oxygen) and oxidative dissolution of AgNPs (assisted by oxygen). In authentic saliva, AgNPs are also shown to be more inert both chemically (silver oxidative dissolution) and electrochemically (electron transfer at an electrode) than in synthetic saliva or aqueous electrolytes. The results thus predict based on the chemical persistence (over a 24-h study) of AgNPs in saliva and hence the minimal release of hazardous Ag+ and reactive oxygen species that the AgNPs are less likely to cause serious harm to the oral cavity but this persistence may enable their transport to other environments.

  • Airborne particulate matter impairs corneal epithelial cells migration via disturbing FAK/RhoA signaling pathway and cytoskeleton organization
    Nanotoxicology (IF 5.811) Pub Date : 2018-02-20
    Yu-Hong Cui, Zi-Xuan Hu, Zi-Xun Gao, Xi-Ling Song, Qing-Yang Feng, Guang Yang, Zhi-Jie Li, Hong-Wei Pan

    Background: Cornea is the outmost structure of the eye and exposed directly to the air pollution. However, little is known about the effect of PM2.5 on corneal epithelium, which is critical for maintenance of cornea homeostasis and visual function.

  • High-throughput tool to discriminate effects of NMs (Cu-NPs, Cu-nanowires, CuNO3, and Cu salt aged): transcriptomics in Enchytraeus crypticus
    Nanotoxicology (IF 5.811) Pub Date : 2018-03-05
    Susana I. L. Gomes, Carlos P. Roca, Natália Pegoraro, Tito Trindade, Janeck J. Scott-Fordsmand, Mónica J. B. Amorim

    The current testing of nanomaterials (NMs) via standard toxicity tests does not cover many of the NMs specificities. One of the recommendations lays on understanding the mechanisms of action, as these can help predicting long-term effects and safe-by-design production. In the present study, we used the high-throughput gene expression tool, developed for Enchytraeus crypticus (4 × 44k Agilent microarray), to study the effects of exposure to several copper (Cu) forms. The Cu treatments included two NMs (spherical and wires) and two copper-salt treatments (CuNO3 spiked and Cu salt field historical contamination). To relate gene expression with higher effect level, testing was done with reproduction effect concentrations (EC20, EC50), using 3 and 7 days as exposure periods. Results showed that time plays a major role in the transcriptomic response, most of it occurring after 3 days. Analysis of gene expression profiles showed that Cu-salt-aged and Cu-nanowires (Nwires) differed from CuNO3 and Cu-nanoparticles (NPs). Functional analysis revealed specific mechanisms: Cu-NPs uniquely affected senescence and cuticle pattern formation, which can result from the contact of the NPs with the worms’ tegument. Cu-Nwires affected reproduction via male gamete generation and hermaphrodite genitalia development. CuNO3 affected neurotransmission and locomotory behavior, both of which can be related with avoidance response. Cu salt-aged uniquely affected phagocytosis and reproductive system development (via different mechanisms than Cu-Nwires). For the first time for Cu (nano)materials, the adverse outcome pathways (AOPs) drafted here provide an overview for common and unique effects per material and linkage with apical effects.

  • RNA sequencing analysis shows that titanium dioxide nanoparticles induce endoplasmic reticulum stress, which has a central role in mediating plasma glucose in mice
    Nanotoxicology (IF 5.811) Pub Date : 2018-03-06
    Hailong Hu, Li Li, Qian Guo, He Zong, Yuheng Yan, Yao Yin, Yu Wang, Yuri Oh, Yujie Feng, Qiong Wu, Ning Gu

    Titanium dioxide nanoparticles (TiO2 NPs) constitute the top five NPs in use today. In this study, oral administration of 50, 100, and 200 mg/kg body weight (b.w.) TiO2 NPs increases plasma glucose in mice, whereas 10 and 20 mg/kg b.w. TiO2 NPs did not. RNA sequencing (RNA-seq) technology was used to investigate genome-wide effects of TiO2 NPs. Clustering analysis of the RNA-seq data showed the most significantly enriched gene ontology terms and KEGG pathways related to the endoplasmic reticulum (ER) and ER stress. Molecular biology verification showed that 50 mg/kg b.w. and higher doses TiO2 NPs activated a xenobiotic biodegradation response and increased expression of cytochrome P450 family genes in mouse livers, thus inducing ER stress in mice. ER stress-activated MAPK and NF-κB pathways and induced an inflammation response, resulting in phosphorylation of the insulin receptor substrate 1 and, consequently, insulin resistance. This was the main mechanism by which TiO2 NPs increased plasma glucose in mice. Meanwhile, ER stress disturbed the monooxygenase system, and thus generated reactive oxygen species (ROS). Relief of ER stress with 4-phenylbutyric acid inhibited all the above effects of TiO2 NPs, including the generation of ROS. Therefore, TiO2 NP-induced ER stress was a decisive factor with a central role in plasma glucose disturbance in mice.

  • Assessment of the in vitro genotoxicity of TiO2 nanoparticles in a regulatory context
    Nanotoxicology (IF 5.811) Pub Date : 2018-03-19
    Sandrine Charles, Stéphane Jomini, Valérie Fessard, Emilie Bigorgne-Vizade, Christophe Rousselle, Cécile Michel

    A review of in vitro genotoxicity studies on titanium dioxide nanoparticles (TiO2-NPs) published between 2010 and 2016 was performed by France in the framework of the CLP Regulation 1272/2008/EC. Neither the few in vivo studies of low quality nor the larger number of acceptable in vitro studies available for genotoxicity allowed France to conclude on the genotoxicity of TiO2-NPs. Based on this work, it was decided to compare the acceptable in vitro studies to understand the reasons for the diverging results observed, such as the materials tested or of the protocols used and their inherent interferences. The systematic review performed on in vitro genotoxicity data for TiO2-NPs was then restricted to studies with the highest level of confidence among studies following OECD guidelines and the largely applied comet assay. Indeed, the aim of this article is to understand why, even if judged of good quality, the 36 publications selected and analyzed did not lead to a clear picture. Some recommendations to be taken into account before performing new in vitro genotoxicity assays for insoluble particles such as TiO2-NPs are proposed. Although secondary genotoxic effects consequent to oxidative stress seem to be the major mechanism responsible for the genotoxicity of TiO2-NPs reported in some studies, primary genotoxic effects cannot be excluded. Further studies are needed to clarify the exact mode of action of TiO2-NPs and to highlight which physicochemical properties lead to their genotoxicity in vitro to ultimately identify a specific combination of parameters that could represent a risk in vivo.

  • Comparative toxicity of three differently shaped carbon nanomaterials on Daphnia magna: does a shape effect exist?
    Nanotoxicology (IF 5.811) Pub Date : 2018-02-01
    Renato Bacchetta, Nadia Santo, Irene Valenti, Daniela Maggioni, Mariangela Longhi, Paolo Tremolada

    The acute toxicity of three differently shaped carbon nanomaterials (CNMs) was studied on Daphnia magna, comparing the induced effects and looking for the toxic mechanisms. We used carbon nano-powder (CNP), with almost spherical primary particle morphology, multi-walled carbon nanotubes (CNTs), tubes of multi-graphitic sheets, and cubic-shaped carbon nanoparticles (CNCs), for which no ecotoxicological data are available so far. Daphnids were exposed to six suspensions (1, 2, 5, 10, 20 and 50 mg L−1) of each CNM, and then microscopically analyzed. Ultrastructural analyses evidenced cellular uptake of nanoparticle in CNP and CNT exposed groups, but not in samples exposed to CNCs. Despite this difference, very similar effects were observed in tissues exposed to the three used CNMs: empty spaces between cells, cell detachment from the basal lamina, many lamellar bodies and autophagy vacuoles. These pathological figures were qualitatively similar among the three groups, but they differed in frequency and severity. CNCs caused the most severe effects, such as partial or complete dissolution of the brush border and thinning of the digestive epithelium. Being the cubic shape not allowed to be internalized into cells, but more effective than others in determining physical damages, we can conclude that shape is an important factor for driving nanoparticle uptake by cells and for determining the acute toxicological endpoints. Shape also plays a key role in determining the kind and the severity of pathologies, which are linked to the physical interactions of CNMs with the exposed tissues.

  • Short-term inhalation study of graphene oxide nanoplates
    Nanotoxicology (IF 5.811) Pub Date : 2018-02-01
    Young Hun Kim, Mi Seong Jo, Jin Kwon Kim, Jae Hoon Shin, Jin Ee Baek, Hye Seon Park, Hyo Jin An, Jong Seong Lee, Boo Wook Kim, Hoi Pin Kim, Kang Ho Ahn, KiSoo Jeon, Seung Min Oh, Ji Hyun Lee, Tomomi Workman, Elaine M. Faustman, Il Je Yu

    Graphene oxides possess unique physicochemical properties with important potential applications in electronics, pharmaceuticals, and medicine. However, the toxicity following inhalation exposure to graphene oxide has not yet been clarified. Therefore, this study conducted a short-term graphene oxide inhalation toxicity analysis using a nose-only inhalation exposure system and male Sprague–Dawley rats. A total of four groups (15 rats per group) were exposed: (1) control (fresh air), (2) low concentration (0.76 ± 0.16 mg/m3), (3) moderate concentration (2.60 ± 0.19 mg/m3), and (4) high concentration (9.78 ± 0.29 mg/m3). The rats were exposed to graphene oxide for 6 h/day for 5 days, followed by recovery for 1, 3, and 21 days. No significant body or organ weight changes were noted after the short-term exposure or during the recovery period. Similarly, no significant systemic effects of toxicological importance were noted in the hematological assays, bronchoalveolar lavage fluid (BAL) inflammatory markers, BAL fluid cytokines, or blood biochemical assays following the graphene oxide exposure or during the post-exposure observation period. Moreover, no significant differences were observed in the BAL cell differentials, such as lymphocytes, macrophages, or polymorphonuclear cells. Graphene oxide-ingested alveolar macrophages as a spontaneous clearance reaction were observed in the lungs of all the concentration groups from post 1 day to post 21 days. Histopathological examination of the liver and kidneys did not reveal any significant test-article-relevant histopathological lesions. Importantly, similar to previously reported graphene inhalation data, this short-term nose-only inhalation study found only minimal or unnoticeable graphene oxide toxicity in the lungs and other organs.

  • Copper oxide nanoparticles induce collagen deposition via TGF-β1/Smad3 signaling in human airway epithelial cells
    Nanotoxicology (IF 5.811) Pub Date : 2018-01-31
    Je-Won Ko, Na-Rae Shin, Ji-Won Park, Sung-Hyeuk Park, In-Chul Lee, Joong-Sun Kim, Jong-Choon Kim, Kyung-Seop Ahn, In-Sik Shin

    Use and application of nanoparticles has increased in recent years. Copper oxide nanoparticles (CuONPs) are one of the most common types of nanoparticles, and they are mainly used as catalysts and preservatives. However, limited toxicity data are available on the toxicity of CuONPs to the respiratory system. We investigated fibrotic responses induced by CuONPs in the respiratory tract and elucidated its underlying mechanism of action in vivo and in vitro experiments. In the mouse model, CuONPs exposure markedly increased transforming growth factor-β1 (TGF-β1) and collagen I expression and Smad3 phosphorylation, combined with elevation of inflammatory mediators including interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α). These alterations were also observed in histological analysis of lung tissue. CuONPs markedly increased inflammatory responses and collagen deposition, accompanied by the elevation of TGF-β1 and collagen I expression in lung tissue. In addition, CuONPs-treated H292 cells showed significantly increased mRNA and protein production of TGF-β1, collagen I, IL-6, and TNF-α; this response was markedly decreased by treatment of a TGF-β1 inhibitor (SB-431542). Taken together, CuONPs induced fibrotic responses in the respiratory tract, closely related to TGF-β1/Smad3 signaling. Therefore, our results raise the necessity of further investigation for the present state of its risk by providing useful information of the toxicity of CuONPs.

  • Metabolomics reveals the depletion of intracellular metabolites in HepG2 cells after treatment with gold nanoparticles
    Nanotoxicology (IF 5.811) Pub Date : 2018-02-02
    Jeremie Zander Lindeque, Alnari Matthyser, Shayne Mason, Roan Louw, Cornelius Johannes Francois Taute

    Studies on the safety of gold nanoparticles (GNPs) are plentiful due to their successful application in drug delivery and treatment of diseases in trials. Cytotoxicity caused by GNPs has been studied on the physiological and biochemical level; yet, the effect of GNPs (particularly gold nano-spheres) on the metabolome of living organisms remains understudied. In this investigation, metabolomics was used to comprehensively study the metabolic alterations in HepG2 cells caused by GNPs; and to investigate the role of representative GNP coatings. GNPs were synthesized, coated and characterized before use on HepG2 cell cultures. Cells were treated for 3 h with citrate-, poly-(sodiumsterene sulfunate)-, and poly-vinylpyrrolidone (PVP)-capped GNPs, respectively. The internalization of the different GNPs and their effect on mitochondrial respiration and the metabolome were studied. Results indicated that the PVP-capped GNPs internalized more and also caused a more observable effect on the metabolome. Conversely, it was the citrate- and poly-(sodiumsterene sulfunate) coated particles that influenced ATP production in addition to the metabolomic changes. A holistic depletion of intracellular metabolites was observed regardless of GNP coating, which hints to the binding of certain metabolites to the particles.

Some contents have been Reproduced with permission of the American Chemical Society.
Some contents have been Reproduced by permission of The Royal Society of Chemistry.
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