Carbon black nanoparticle instillation induces sustained inflammation and genotoxicity in mouse lung and liver

Health Canada, Environmental and Radiation Health Sciences Directorate, Mechanistic Studies Division, Tunney's Pasture, Ottawa, Canada.
Particle and Fibre Toxicology (Impact Factor: 7.11). 02/2012; 9(1):5. DOI: 10.1186/1743-8977-9-5
Source: PubMed


Widespread occupational exposure to carbon black nanoparticles (CBNPs) raises concerns over their safety. CBNPs are genotoxic in vitro but less is known about their genotoxicity in various organs in vivo.
We investigated inflammatory and acute phase responses, DNA strand breaks (SB) and oxidatively damaged DNA in C57BL/6 mice 1, 3 and 28 days after a single instillation of 0.018, 0.054 or 0.162 mg Printex 90 CBNPs, alongside sham controls. Bronchoalveolar lavage (BAL) fluid was analyzed for cellular composition. SB in BAL cells, whole lung and liver were assessed using the alkaline comet assay. Formamidopyrimidine DNA glycosylase (FPG) sensitive sites were assessed as an indicator of oxidatively damaged DNA. Pulmonary and hepatic acute phase response was evaluated by Saa3 mRNA real-time quantitative PCR.
Inflammation was strongest 1 and 3 days post-exposure, and remained elevated for the two highest doses (i.e., 0.054 and 0.162 mg) 28 days post-exposure (P < 0.001). SB were detected in lung at all doses on post-exposure day 1 (P < 0.001) and remained elevated at the two highest doses until day 28 (P < 0.05). BAL cell DNA SB were elevated relative to controls at least at the highest dose on all post-exposure days (P < 0.05). The level of FPG sensitive sites in lung was increased throughout with significant increases occurring on post-exposure days 1 and 3, in comparison to controls (P < 0.001-0.05). SB in liver were detected on post-exposure days 1 (P < 0.001) and 28 (P < 0.001). Polymorphonuclear (PMN) cell counts in BAL correlated strongly with FPG sensitive sites in lung (r = 0.88, P < 0.001), whereas no such correlation was observed with SB (r = 0.52, P = 0.08). CBNP increased the expression of Saa3 mRNA in lung tissue on day 1 (all doses), 3 (all doses) and 28 (0.054 and 0.162 mg), but not in liver.
Deposition of CBNPs in lung induces inflammatory and genotoxic effects in mouse lung that persist considerably after the initial exposure. Our results demonstrate that CBNPs may cause genotoxicity both in the primary exposed tissue, lung and BAL cells, and in a secondary tissue, the liver.

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Available from: Carole Yauk, Oct 03, 2015
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    • "This was followed by the main study were groups of mice were given 0, 2, 6 or 18 mg ZnO NP or 162 mg carbon black Printex 90 by a single intratracheal instillation. Printex 90 is a highly inflammatory particle that has been extensively examined and used as a benchmark particle (Jacobsen et al., 2007, 2008, 2011; Jackson et al., 2012; Bourdon et al., 2012; Hogsberg et al., 2013; Vesterdal et al., 2010; Kyjovska et al., 2015a). Each exposed group consisted of six mice (N ¼ 6) and each control group of twelve mice (N ¼ 12) and was planned to be killed after 1d and 3d. "
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    ABSTRACT: Inhalation is the main pathway of ZnO exposure in the occupational environment but only few studies have addressed toxic effects after pulmonary exposure to ZnO nanoparticles (NP). Here we present results from three studies of pulmonary exposure and toxicity of ZnO NP in mice. The studies were prematurely terminated because interim results unexpectedly showed severe pulmonary toxicity. High bolus doses of ZnO NP (25 up to 100 mg; �1.4 mg/kg) were clearly associated with a dose dependent mortality in the mice. Lower doses (�6 mg; �0.3 mg/kg) elicited acute toxicity in terms of reduced weight gain, desquamation of epithelial cells with concomitantly increased barrier permeability of the alveolar/ blood as well as DNA damage. Oxidative stress was shown via a strong increase in lipid peroxidation and reduced glutathione in the pulmonary tissue. Two months post-exposure revealed no obvious toxicity for 12.5 and 25 mg on a range of parameters. However, mice that survived a dose (50 mg; 2.7 mg/kg) had an increased pulmonary collagen accumulation (fibrosis) at a similar level as a high bolus dose of crystalline silica. The recovery from these toxicological effects appeared dose-dependent. The results indicate that alveolar deposition of ZnO NP may cause significant adverse health effects.
    Food and Chemical Toxicology 08/2015; Food Chem. Toxicol.. · 2.90 Impact Factor
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    • "CB particles (Printex 90) were obtained from Evonik Industries, Frankfurt, Germany (primary particle size 14 nm; surface area 300 m2/g). Printex 90 is an extensively studied model NP and has been characterized elsewhere [20], [21]. The mean size of the particles suspended in media was 85±38 nm [13]. "
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    ABSTRACT: Exposure to nanoparticles (NPs) may cause vascular effects including endothelial dysfunction and foam cell formation, with oxidative stress and inflammation as supposed central mechanisms. We investigated oxidative stress, endothelial dysfunction and lipid accumulation caused by nano-sized carbon black (CB) exposure in cultured human umbilical vein endothelial cells (HUVECs), THP-1 (monocytes) and THP-1 derived macrophages (THP-1a). The proliferation of HUVECs or co-cultures of HUVECs and THP-1 cells were unaffected by CB exposure, whereas there was increased cytotoxicity, assessed by the LDH and WST-1 assays, especially in THP-1 and THP-1a cells. The CB exposure decreased the glutathione (GSH) content in THP-1 and THP-1a cells, whereas GSH was increased in HUVECs. The reactive oxygen species (ROS) production was increased in all cell types after CB exposure. A reduction of the intracellular GSH concentration by buthionine sulfoximine (BSO) pre-treatment further increased the CB-induced ROS production in THP-1 cells and HUVECs. The expression of adhesion molecules ICAM-1 and VCAM-1, but not adhesion of THP-1 to HUVECs or culture dishes, was elevated by CB exposure, whereas these effects were unaffected by BSO pre-treatment. qRT-PCR showed increased VCAM1 expression, but no change in GCLM and HMOX1 expression in CB-exposed HUVECs. Pre-exposure to CB induced lipid accumulation in THP-1a cells, which was not affected by the presence of the antioxidant N-acetylcysteine. In addition, the concentrations of CB to induce lipid accumulation were lower than the concentrations to promote intracellular ROS production in THP-1a cells. In conclusion, exposure to nano-sized CB induced endothelial dysfunction and foam cell formation, which was not dependent on intracellular ROS production.
    PLoS ONE 09/2014; 9(9):e106711. DOI:10.1371/journal.pone.0106711 · 3.23 Impact Factor
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    • "ion , there was no difference in neutrophil influx between vehicle control animals and animals instilled with the three lower doses of NPCB . The exposure also changed the distribution of macro - phages , eosinophils , and lymphocytes . The change in the number of macrophages after instillation of 162 mg NPCB was similar to previous observations [ Bourdon et al . , 2012 ] . The number of lymphocytes in BAL fluid was stat - istically significantly increased 3 days post - exposure to 0 . 67 mg NPCB ( vehicle controls : 0 ."
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    ABSTRACT: We previously observed genotoxic effects of carbon black nanoparticles at low doses relative to the Danish Occupational Exposure Limit (3.5 mg/m3). Furthermore, DNA damage occurred in broncho-alveolar lavage (BAL) cells in the absence of inflammation, indicating that inflammation is not required for the genotoxic effects of carbon black. In this study, we investigated inflammatory and acute phase response in addition to genotoxic effects occurring following exposure to nanoparticulate carbon black (NPCB) at even lower doses. C57BL/6JBomTac mice were examined 1, 3, and 28 days after a single instillation of 0.67, 2, 6, and 162 µg Printex 90 NPCB and vehicle. Cellular composition and protein concentration was evaluated in BAL fluid as markers of inflammatory response and cell damage. DNA strand breaks in BAL cells, lung, and liver tissue were assessed using the alkaline comet assay. The pulmonary acute phase response was analyzed by Saa3 mRNA real-time quantitative PCR. Instillation of the low doses of NPCB induced a slight neutrophil influx one day after exposure. Pulmonary exposure to small doses of NPCB caused an increase in DNA strand breaks in BAL cells and lung tissue measured using the comet assay. We interpret the increased DNA strand breaks occurring following these low exposure doses of NPCB as DNA damage caused by primary genotoxicity in the absence of substantial inflammation, cell damage, and acute phase response. Environ. Mol. Mutagen. 2014. © 2014 The Authors Environmental and Molecular Mutagenesis published by Wiley Periodicals, Inc. on behalf of Environmental Mutagen Society
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