GENERATION OF REACTIVE OXYGEN SPECIES BY X-RAY IRRADIATED SILICON NANOPARTICLES AND ITS RELEVANCE IN TUMOR CELL SURVIVAL.

DAVID GARA, PEDRO; GARABANO, NATALIA; M.J. LLANSOLA PORTOLÉS; RUBERT, ALDO; D.O. MÁRTIRE.; O.R. CASAS; SANCHEZ, V.N.; GONZALEZ, MÓNICA C.; MÓNICA L. KOTLER

La Serena, Chile

Congreso; X Encuentro Latinoamericano de Fotoquímica y Fotobiología; 2010

Universidad de Santiago de Chile

Silicon nanoparticles (Si-NP) of 1-10 nm size show strong photoluminescence due to quantum confinement, being wavelength emission dependent on size and surface coating. Their strong luminescence and photostability make them excellent candidates as specific markers in biological environments for “in vitro” and “in vivo” applications. In the present work we study the incorporation of Si-NP to tumor cells, its cytotoxicity and the effect of ionizing radiation on the Si-NP properties and reactive oxygen species (ROS) generation. Aqueous suspensions of derivatized and naked Si-NP were irradiated with X-rays (electron linear accelerator Varian Clinac, 4 MeV). The absorbed doses were similar to those used in patients with a standard fractionation scheme. Irradiation increased oxidation and decreased the size of Si-NP. The ability of NP-Si to generate reactive ROS and its steady-state concentration were determined following the consumption of specific probes by HPLC. It was observed that ROS generation was of a higher magnitude order in the presence than in the absence of NP-Si. The incorporation of naked and derivatized NP-Si to rat glioma C6 cells was demonstrated by microscopy and spectrofluorometry at different incubations times. Subcellular localization of Si-NP was determined by using fluorescent dyes that specifically stain mitochondria (Mitotracker Red CMXRos) and lysosomes (LYSO Tracker Red DND99). Both types of NP-Si were early localized in lysosomas. Si-NP cytotoxicity was determined both in the presence and absence of ionizing radiation by morphological analysis and cell viability measurement (Neutral Red, Crystal Violet and LDH). The morphological evaluation showed signs of cellular stress with necrotic characteristics at 50μg/ml, in agreement with viability tests. When cells with Si-NP were irradiated with 1Gy, the decrease in cell viability was approximately 40% (20% higher than the corresponding to 2Gy). The ability of Si-NP incorporated into C6 cells to generate ROS by irradiation was studied employing the DCFH-DA probe. It was observed that ROS generation is a function of both, Si-NP concentration and irradiation doses.