Toxicology Letters 209 (2012) 136–145 Contents lists available at SciVerse ScienceDirect Toxicology Letters jou rn al h om epa ge: www.elsevier.com/locate/toxlet Gene expression profiling of A549 cells exposed to Milan PM2.5 Maurizio Gualtieri a,∗ , Eleonora Longhin a , Michela Mattioli b,1 , Paride Mantecca a , Valentina Tinaglia b , Eleonora Mangano c , Maria Carla Proverbio b , Giuseppina Bestetti a , Marina Camatini a , Cristina Battaglia b a POLARIS Research Center, Department of Environmental Science, University of Milano Bicocca, 1, piazza della Scienza, I-20126 Milan, Italy b Dipartimento di Scienze e Tecnologie biomediche and Scuola di dottorato di medicina molecolare, Università degli Studi di Milano, 93, via F.lli Cervi, I-20090 Segrate, Italy c Institute of Biomedical Technology (ITB), CNR, 93, via F.lli Cervi, I-20090 Segrate, Italy a r t i c l e i n f o Article history: Received 13 September 2011 Received in revised form 17 November 2011 Accepted 18 November 2011 Available online 9 December 2011 Keywords: PM2.5 Gene expression DNA damage A549 ROS a b s t r a c t Background: Particulate matter (PM) has been associated to adverse health effects in exposed population and DNA damage has been extensively reported in in vitro systems exposed to fine PM (PM2.5). The ability to induce gene expression profile modulation, production of reactive oxygen species (ROS) and strand breaks to DNA molecules has been investigated in A549 cells exposed to winter and summer Milan PM2.5. Results: A549 cells, exposed to 10 g/cm 2 of both winter and summer PM2.5, showed increased cyto- toxicity at 24 h and a significant increase of ROS at 3 h of treatment. Despite these similar effects winter PM induced a higher number of gene modulation in comparison with summer PM. Both PMs modulated genes related to the response to xenobiotic stimuli (CYP1A1, CYP1B1, TIPARP, ALDH1A3, AHRR) and to the cell–cell signalling (GREM1) pathways with winter PM2.5 inducing higher fold increases. Moreover the winter fraction modulated also JUN (cell–cell signalling), GDF15, SIPA1L2 (signal transduction), and HMOX1 (oxidative stress). Two genes, epiregulin (EREG) and FOS-like antigen1 (FOSL1), were significantly up-regulated by summer PM2.5. The results obtained with the microarray approach have been confirmed by qPCR and by the analysis of CYP1B1 expression. Comet assay evidenced that winter PM2.5 induced more DNA strand breaks than the summer one. Conclusion: Winter PM2.5 is able to induce gene expression alteration, ROS production and DNA damage. These effects are likely to be related to the CYP enzyme activation in response to the polycyclic aromatic hydrocarbons (PAHs) adsorbed on particle surface. © 2011 Elsevier Ireland Ltd. All rights reserved. 1. Introduction The Po Valley is a densely populated and industrialized area, and one of the most PM-polluted zones in Europe (Koelemeijer et al., 2006). According to emission inventory data for the Milan municipality, about 65% of the annual PM2.5 emissions derive from the traffic source, 20% from combustion for house heating and only the remaining 15% from industrial emissions, since heavy industries are no longer operating in the city area (Lombardy Region, 2007). Comparing the annual average of PM2.5 concen- tration found in this study and the previous ones (Lonati et al., 2008; Marcazzan et al., 2001) with the levels of other European countries (Querol et al., 2004), it results that annual PM2.5 levels ∗ Corresponding author. Tel.: +39 02 6448 2928; fax: +39 02 6448 2996. E-mail address: maurizio.gualtieri@unimib.it (M. Gualtieri). 1 Current address: Therasis, Inc., 462 First Avenue, New York, NY 10016, United States. in Milan (34 g/m 3 ) are far higher than those observed at urban background sites in Northern and Southern Europe (8–15 g/m 3 and 19–25 g/m 3 , respectively), and slightly higher than those observed in Central Europe (16–30 g/m 3 ). On the other hand it has been reported that the PM2.5 levels in Milan are comparable to those of traffic exposed sites in Central and Southern Europe (22–39 g/m 3 and 28–35 g/m 3 , respectively). Therefore, as PM2.5 chemical composition is similar to that of other European cities, rel- atively high PM2.5 concentration levels are peculiar of the Milan area. The Milan PM2.5 fraction is dominated by combustion derived particles, consisting of a carbonaceous core with organic and inor- ganic compounds adsorbed on its surface (Sharma et al., 2007; Sevastyanova et al., 2008; Zerbi et al., 2008; Gualtieri et al., 2009) and its chemical characterization has received attention since 1997 (Marcazzan et al., 2001; Lonati et al., 2007, 2008). The morphological damages produced on the human alveo- lar epithelial cells (A549) exposed to Milan winter PM2.5 have been previously reported (Gualtieri et al., 2009) as well as the 0378-4274/$ – see front matter © 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.toxlet.2011.11.015