Toxicology Letters 207 (2011) 128–136 Contents lists available at SciVerse ScienceDirect Toxicology Letters jou rn al h om epa ge: www.elsevier.com/locate/toxlet Cellular distribution and degradation of cobalt ferrite nanoparticles in Balb/3T3 mouse fibroblasts Patrick Marmorato a , Giacomo Ceccone a,∗ , Alessandra Gianoncelli b , Lorella Pascolo b , Jessica Ponti a , Franc ¸ ois Rossi a , Murielle Salomé c , Burkhard Kaulich b , Maya Kiskinova b a European Commission, Joint Research Centre, Institute for Health and Consumer Protection, Via E. Fermi 2749, 21027 Ispra (VA), Italy b ELETTRA, Sincrotrone Trieste, Strada Statale 149, 34149 Basovizza, Trieste, Italy c European Synchrotron Radiation Facility (ESRF), 6 rue Jules Horowitz, 38000 Grenoble, France a r t i c l e i n f o Article history: Received 18 April 2011 Received in revised form 30 August 2011 Accepted 31 August 2011 Available online 8 September 2011 Keywords: X-ray microscopy X-ray fluorescence Fibroblasts Magnetic nanoparticles a b s t r a c t The effect of the concentration of cobalt ferrite (CoFe 2 O 4 ) nanoparticles (NPs) on their intracellular loca- tion and distribution has been explored by synchrotron radiation X-ray and fluorescence microscopy (SR-XRF) monitoring the evolution of NPs elemental composition as well. In cells exposed to low con- centrations of CoFe 2 O 4 NPs, the NPs preferentially segregate in the perinuclear region preserving their initial chemical content. At concentrations exceeding 500 M the XRF spectra indicate the presence of Co and Fe also in the nuclear region, accompanied by sensible changes in the cellular morphology. The increase of the Co/Fe ratio measured in the nuclear compartment indicates that above certain concen- trations the CoFe 2 O 4 NPs intracellular distribution could be accompanied by biodegradation resulting in Co accumulation in the nucleus. © 2011 Elsevier Ireland Ltd. All rights reserved. 1. Introduction Rapid growth of biomedical applications of NPs as drug-delivery agents, biosensors or imaging contrast agents imposes urgent request for better understanding of their intracellular fate and interactions in order to prove their biocompatibility. One of the crucial problems is that reducing the particle size dramatically improves the bioavailability of gene or drug delivery systems but also enhances the particle reactivity, which poses potential health hazards. Although nanotechnology provides means to overcome the enhanced reactivity by proper surface passivation (Mirkin and Andrew, 2000) it cannot prevent biodegradation in the cellular environment and predict the cellular responses induced by the degraded NPs. For example, biodegraded NPs may accumulate within cells and lead to intracellular changes such as disruption of organelle integrity or gene alterations. Therefore understanding the properties of NPs and their effect is crucial before any clinical use, which requires multidisciplinary efforts for developing a proper methodology by implementing complementary tools for charac- terization of the NPs physico-chemical status and the relevant biochemical processes occurring in the cells (Lewinski et al., 2008; Nel et al., 2009; Ortega et al., 2009a). Amongst the number of dif- ferent imaging and microscopy techniques, synchrotron radiation ∗ Corresponding author. Tel.: +39 033785475; fax: +39 0332785787. E-mail address: giacomo.ceccone@jrc.ec.europa.eu (G. Ceccone). X-ray imaging in combination with X-ray fluorescence (SR-XRF) and/or X-ray Absorption Near Edge (XANES) micro-spectroscopy is very attractive. These techniques can map the distribution of the NPs in the cells and quantify their actual chemical compo- sition with submicron resolution (Paunesku et al., 2006; Fahrni, 2007) and have already been successfully used for environmen- tal analysis of single cells (Zhao and Le, 2007), cell differentiation (Ide-Ektessabi, 2007a) and trace element analysis in biological sam- ples/tissues (Kwiatek et al., 2001; Carvalho et al., 2007; Yoshida et al., 2003; Szczerbowska-Boruchowska et al., 2003). Moreover, SR-XRF has been used, in combination with nuclear techniques, to detect, map and quantify toxic elements such as chromium and cobalt in cells (Ortega et al., 2003). Up to now the SR-XRF mapping of cells with NPs has exclusively been used for determining their intracellular distribution, e.g. for TiO 2 -DNA oligonucleotide nanoconiugates (Paunesku et al., 2007), quantum dots in human cells (Corezzi et al., 2009) and carbon nan- otubes in macrophages (Bussy et al., 2008). However, to the best of our knowledge, there are no reports about possible changes in the chemistry of NPs penetrating in different cell compartments, as reported in the present study for CoFe 2 O 4 NPs up-taken by Balb/3T3 mouse fibroblasts. CoFe 2 O 4 NPs are super-paramagnetic NPs with possible future biomedical applications (Baldi et al., 2007a; Mornet et al., 2006) spanning from cell separation, purification and contrast agents for magnetic resonance imaging (MRI) (Joshi et al., 2009) to drug deliv- ery (Arruebo et al., 2007), biosensors (Chemla et al., 2000), and 0378-4274/$ – see front matter © 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.toxlet.2011.08.026