S192 Abstracts / Toxicology Letters 229S (2014) S40–S252 intestine barriers (co-culture) and there is surfactant secretion in the tetraculture system at the air-liquid interface (ALI) and mucus production in the intestinal coculture. The tetraculture model was exposed to different realis- tic amounts of diesel exhaust particulate matter (80 ng/cm 2 , or 240 ng/cm 2 ). A clear dose-dependent translocation of the transcription factor Nrf2, which regulates gene expression in response to oxidative stress, was observed after 4 h of incubation in the endothelial cells without reduction of cell viability. The alveolar model is able to detect secondary induced toxicity upon realistic exposure to environmental relevant concentration of particulate matter. The intestinal coculture model was used to evaluate effects of Ag 20 and 200 nm particles on the metabolic activity, oxidative stress and pro-inflammatory cytokine release. AgNO 3 induced a reduc- tion in metabolic activity in a dose dependent manner whereas no reduction was observed for both Ag particles. Ag was found to be homogenously distributed in the cell with aggregates observed for Ag 20 with a 5-fold increase in IL8 release. The proteomic data revealed that both Ag particles induced oxidative stress pathways and affected cytoskeleton, but regulated different sets of proteins compared to AgNO 3 . Overall these two systems may become valuable tools for toxi- cological studies. http://dx.doi.org/10.1016/j.toxlet.2014.06.650 P-3.138 Protein carbonylation as a marker of oxidative stress induced by nanoparticles: Analysis of 16 inorganic nanoparticles Marc D. Driessen 1 , Rainer Ossig 2 , Jürgen Schnekenburger 2 , Antje Vennemann 3 , Martin Wiemann 3 , Andreas Luch 1 , Andrea Haase 1,* 1 German Federal Institute for Risk Assessment (BfR), Berlin, Germany, 2 Biomedical Technology Center, Westfälische Wilhelms-Universität, Muenster, Germany, 3 IBE R&D gGmbH, Muenster, Germany The BMBF-funded project “nanoGEM” follows a systematic approach to understand hazards associated with different types of nanoparticles (NP). Oxidative stress is considered to be a major paradigm to explain NP toxicity. Here we focused on protein car- bonylation as a consequence of oxidative stress for a set of 16 different nanoparticles, used as either plane materials or with dif- ferent surface coatings. In parallel several in vitro and in vivo toxicity endpoints have been analyzed. We used NP of 10 nm (ZrO 2 ), 15 nm (SiO 2 ) and 50 nm or 200 nm (Ag), furnished either with acidic, basic or polymeric functionali- ties and TiO 2 , ZnO, BaSO 4 and AlOOH as references. In a screening approach we studied time- and dose-dependent carbonylation of all 16 NP in NRK-52E cells via 1D immunoblots. Data were cor- related with cytotoxicity (WST-8, LDH assay) and ROS formation (DCFDA assay). Furthermore we applied a 2D proteomics approach combined with MALDI-MS/MS to identify the proteins modified. Finally, for several NP we analyzed lung tissues after in vivo instil- lation in rats. Eight out of 16 NP induced protein carbonylation in NRK-52E cells. Observed protein carbonylation correlated well with overall toxicity. The 2D approach revealed a complex and distinct pattern of carbonyls. Modified proteins were identified as cytoskeleton pro- teins, heat shock proteins or proteins of major cellular pathways (i.e. glycolysis). We also observed carbonyl modifications in lung tissue homogenates of rats intratracheally instilled with the same NP. Taken together, analysis of protein carbonylation is a useful tool for the analysis and mechanistic understanding of ROS dependent NM toxicity. http://dx.doi.org/10.1016/j.toxlet.2014.06.651 P-3.139 Nanoparticle-induced oxidative stress alters phospho-tyrosine patterns in mammalian cells: Results of an SH2 profiling approach Marc D. Driessen 1 , Rainer Ossig 2 , Jürgen Schnekenburger 2 , Antje Vennemann 3 , Martin Wiemann 3 , Andreas Luch 1 , Peter Nollau 4 , Andrea Haase 1,* 1 German Federal Institute for Risk Assessment (BfR), Berlin, Germany, 2 Biomedical Technology Center, Westfälische Wilhelms-Universität, Muenster, Germany, 3 IBE R&D gGmbH, Muenster, Germany, 4 Forschungsinstitut Kinderkrebs-Zentrum Hamburg, University Medical Center Hamburg-Eppendorf, Hamburg, Germany The BMBF-funded project “nanoGEM” follows a systematic approach to understand hazards associated with different types of nanoparticles (NP). Oxidative stress is considered to be a major paradigm to explain NP toxicity and can result in altered levels of tyrosine phosphorylation, probably by protein tyrosine phos- phatase inhibition which are major components of various cellular signaling pathways. Here we focused on global changes of tyrosine phosphorylation resulting from treatment with 16 NP bearing dif- ferent surface coatings. In parallel several well established in vitro and in vivo toxicity endpoints were analyzed. We used ZrO 2 , SiO 2 Ag NP, either plane or acidic, basic or polymeric functionalisation and in addition TiO 2 , ZnO, BaSO 4 and AlOOH as references. We analyzed NP treated NRK-52E cells by far western blot analysis with a set of 70 src homolgy 2 (SH2) domains known to differentially bind phospho-tyrosine sites. From these 70 SH2 domains, a set of 9 domains was selected and cellular reactions after SiO 2 and Ag NP treatment were studied in detail. Further- more we used these 9 SH2 domains to study changes in signaling pathways in lung tissue lysates after in vivo instillation in rats. Most NP caused alterations in phospho-tyrosine dependent cell signaling. Strongest responses were observed for cytotoxic NP (SiO 2 naked, ZnO), fitting very well to overall observed toxicity, additionally enabling us to detect subtle changes in phospho- tyrosine dependent signaling also for non-toxic NP. SH2 profiling appears to be a powerful tool to study cellular responses after NP treatment and opens insights into signaling mechanisms underlying toxicty. http://dx.doi.org/10.1016/j.toxlet.2014.06.652 P-3.140 Critical role of interleukin 1 for the initiation and resolution of pulmonary inflammation induced by carbon nanomaterials Nunja C. Habel 1,* , David Kutschke 1 , Oliver Eickelberg 1,2 , Silke Meiners 1 , Tobias Stoeger 1 1 Comprehensive Pneumology Center, Institute of Lung Biology and Disease, Helmholtz Zentrum München, Munich, Germany, 2 Institute of Experimental Pneumology, Munich, Germany Inhalation of carbon nanoparticles has been shown to cause acute lung inflammation, and particle shape has been related to inflammasome and downstream Il1-R1 receptor signaling