Cytotoxicity and genotoxicity induced in vitro by solvent-extractable organic matter of size-segregated urban particulate matter * Ekaterini Velali a, 1 , Eleni Papachristou a, 1 , Anastasia Pantazaki a, ** , Theodora Choli-Papadopoulou a , Nikoleta Argyrou b, 1 , Theodora Tsourouktsoglou b, 1 , Stergios Lialiaris b, 1 , Alexandros Constantinidis b, 1 , Dimitrios Lykidis b, 1 , Thedore S. Lialiaris b , Athanasios Besis c, 1 , Dimitra Voutsa c , Constantini Samara c, * a Laboratory of Biochemistry, Department of Chemistry, Aristotle University of Thessaloniki, GR-54124 Thessaloniki, Greece b Demokrition University of Thrace, Faculty of Medicine, Department of Genetics, Alexandroupolis 68100, Greece c Environmental Pollution Control Laboratory, Department of Chemistry, Aristotle University of Thessaloniki, GR-54124 Thessaloniki, Greece article info Article history: Received 24 April 2016 Received in revised form 21 August 2016 Accepted 2 September 2016 Available online 6 September 2016 Keywords: 8-OHdG Comet assay DNA MTT assay PARP SCE assay TNF-a abstract Three organic fractions of different polarity, including a non polar organic fraction (NPOF), a moderately polar organic fraction (MPOF), and a polar organic fraction (POF) were obtained from size-segregated (<0.49, 0.49e0.97, 0.97e3 and >3 mm) urban particulate matter (PM) samples, and tested for cytotox- icity and genotoxicity using a battery of in vitro assays. The cytotoxicity induced by the organic PM fractions was measured by the mitochondrial dehydrogenase (MTT) cell viability assay applied on MRC-5 human lung epithelial cells. DNA damages were evaluated through the comet assay, determination of the poly(ADP-Ribose) polymerase (PARP) activity, and the oxidative DNA adduct 8-hydroxy-deoxyguanosine (8-OHdG) formation, while pro-inflammatory effects were assessed by determination of the tumor ne- crosis factor-alpha (TNF-a) mediator release. In addition, the Sister Chromatid Exchange (SCE) induc- ibility of the solvent-extractable organic matter was measured on human peripheral lymphocyte. Variations of responses were assessed in relation to the polarity (hence the expected composition) of the organic PM fractions, particle size, locality, and season. Organic PM fractions were found to induce rather comparable Cytotoxicity and genotoxicity of PM appeared to be rather independent from the polarity of the extractable organic PM matter (EOM) with POF often being relatively more toxic than NPOF or MPOF. All assays indicated stronger mass-normalized bioactivity for fine than coarse particles peaking in the 0.97e3 and/or the 0.49e0.97 mm size ranges. Nevertheless, the air volume-normalized bioactivity in all assays was highest for the <0.49 mm size range highlighting the important human health risk posed by the inhalation of these quasi-ultrafine particles. © 2016 Elsevier Ltd. All rights reserved. 1. Introduction Ambient particulate matter (PM) is associated with short-term and long-term health effects (hospital admissions, premature mortality, morbidity, lung cancer, cardiovascular and cardiopul- monary diseases, etc) (Pope et al., 2009). Toxicological studies have shown that PM has several mecha- nisms of adverse cellular effects, such as cytotoxicity, oxidative stress, generation of reactive oxygen species (ROS), DNA damage, mutagenicity, and stimulation of pro-inflammatory factors (de Kok et al., 2006; Valavanidis et al., 2008, 2009; Falcon-Rodriguez et al., 2016), however a complete understanding of the mechanisms of action at cell level still lacks. PM size and composition are critical characteristics determining it's biological effects. Many studies have shown that the potential to elicit biological effects are stron- ger for fine and ultrafine particles (UFPs) because they can pene- trate deeper into the respiratory tract reaching the alveoli, moreover because these particles are more enriched with toxic and carcinogenic compounds than coarse particles (Topinka et al., * This paper has been recommended for acceptance by Dr. Chen Da. * Corresponding author. ** Corresponding author. E-mail addresses: natasa@chem.auth.gr (A. Pantazaki), csamara@chem.auth.gr (C. Samara). 1 Young researchers with equal contribution to this work. Contents lists available at ScienceDirect Environmental Pollution journal homepage: www.elsevier.com/locate/envpol http://dx.doi.org/10.1016/j.envpol.2016.09.001 0269-7491/© 2016 Elsevier Ltd. All rights reserved. Environmental Pollution 218 (2016) 1350e1362