Biochemical and cellular effects of electrophiles present in ambient air samples Noriko Iwamoto a , Akira Nishiyama a , Arantzazu Eiguren-Fernandez b, c , William Hinds b, c , Yoshito Kumagai a, b , John R. Froines b, c , Arthur K. Cho b, c , Masaru Shinyashiki b, c, * a Doctoral Programs in Medical Sciences, Graduate School of Comprehensive Human Sciences, University of Tsukuba,1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan b Southern California Particle Center, University of California Los Angeles, Los Angeles, CA 90095, USA c Department of Environmental Health Sciences, UCLA School of Public Health, Los Angeles, CA 90095, USA article info Article history: Received 29 October 2009 Received in revised form 28 January 2010 Accepted 29 January 2010 Keywords: Ambient vapor-phase Electrophiles Particulate matters Thiol proteins Cell responses abstract Ambient vapor-phase samples collected in Riverside, California had shown that both redox and elec- trophilic activity were present, with the vapor phase containing higher levels of electrophiles than the particle phase. In this study, the biochemical effects of the vapor-phase electrophiles were examined using the purified thiol proteins, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), protein tyrosine phosphatase 1B (PTP1B) and KELCH-1 like ECH-associated protein 1 (Keap1). The results demonstrated that the thiol proteins were inactivated by the vapor-phase samples through covalent modifications. Next, two cellular responses, epidermal growth factor receptor (EGFR)/mitogen-activated protein (MAP) kinase and NF-E2-related factor 2 (Nrf2), to the ambient vapor-phase samples were assessed in A549 and RAW 264.7 cell lines, respectively. The vapor-phase samples, at non-oxidative concentrations, increased phosphorylation of EGFR, which is negatively regulated by PTP1B, and its downstream MAP kinase, extracellular signal-regulated kinase (ERK)1/2. Activation of Nrf2, which requires Keap1 alkylation, and expression of its downstream proteins were also observed. The electrophilic compounds present in ambient vapor-phase were shown to modify cellular proteins through covalent modification and to activate diverse cellular responses that can lead to inflammatory and adaptive responses. Ó 2010 Elsevier Ltd. All rights reserved. 1. Introduction Numerous studies (Frampton et al., 1999; Verones and Oortgiesen, 2001; Veronesi et al., 2002; Calcabrini et al., 2004; Donaldson et al., 1997; Billet et al., 2007; Rumelhard et al., 2007) have described the toxicological properties of airborne particulate matter (PM), but little attention has been paid to the volatile components of ambient air that could initiate toxicological changes upon exposure. The participation of vapor-phase components of ambient air in the toxicity of air samples has been demonstrated in the studies using freshly emitted gasoline and diesel engine exhaust (Lund et al., 2007; Seagrave et al., 2003). The semi-volatile organic compounds were found to be more toxic than those in the particles. This laboratory has been studying the chemical properties of PM as they relate to two common mechanisms of toxicity, the ability to generate reactive oxygen species (ROS) (Cho et al., 2005) and the ability to form covalent bonds (Shinyashiki et al., 2008, 2009). As part of an extended study of air samples from sites in the Los Angeles Basin (LAB), we have initially conducted collection campaigns in Riverside, California, a so called receptor site in which an air mass generated in the west side of the LAB is modified by components in the atmosphere and by light to undergo chemical changes (Eiguren-Fernandez et al., in press). In these collections, we were interested in collecting particles and vapors simultaneously. The results of the chemical analyses showed electrophiles to be present in both the particle and vapor-phase, but the electrophilic activity in the vapor-phase was 15-fold higher than that in the particle phase (Eiguren-Fernandez et al., in press). As electrophiles form covalent bonds with cellular nucleophiles, their actions could be long lasting causing different responses from those resulting from increases in cellular ROS, which can be reduced by cellular antioxidant mechanisms. In prior studies, the electrophilic quinone, 1,2-naphthoquinone (1,2-NQ), and protein tyrosine phosphatase 1B (PTP1B) were found to undergo a Michael addition reaction resulting in covalent bond formation (Iwamoto et al., 2007). PTP1B is a negative regulator of the epidermal growth factor receptor (EGFR) (McCole et al., 2007), a receptor tyrosine kinase associated with multiple cellular responses including apoptosis, inflammation and cell proliferation * Corresponding author at: Department of Environmental Health Sciences, UCLA School of Public Health, 650 Charles E. Young Drive South, Los Angeles, CA 90095, USA. Tel.: þ1 310 794 4178; fax: þ1 310 206 9903. E-mail address: masaru@ucla.edu (M. Shinyashiki). Contents lists available at ScienceDirect Atmospheric Environment journal homepage: www.elsevier.com/locate/atmosenv 1352-2310/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.atmosenv.2010.01.045 Atmospheric Environment 44 (2010) 1483e1489