IRINEL ANCA RADU, MIHAELA MOISESCU, DANIELA IANCU, T. SAVOPOL, EUGENIA KOVÁCS
Department of Biophysics and Cell Biotechnology, “Carol Davila” University of Medicine and Pharmacologie, P.O.Box 35-43, Bucharest, Romania
Abstract. Our study aims to assess the effect of photodynamic therapy (PDT) using ALA (5-aminolaevulinic acid, a precursor of intracellular porphyrins) and red LASER light on Human Caucasian dysplastic oral keratinocytes (DOK). As a first step we determined the best experimental conditions and the amount of reactive oxygen species (ROS) produced as a result of ALA-PDT. DOK cells were incubated with various concentrations of ALA, for different periods of time, in order to determine the optimal concentration of the photosensitizer and duration of incubation. The intracellular amount of ALA-induced protoporphyrin IX (PpIX) was estimated by its fluorescence emission spectra. The photolysis of PpIX in cells irradiated with a red laser was measured by a decrease in the intensity of these spectra. Intracellular ROS were evaluated using a fluorescent nonselective marker, 2’,7’–dichlorofluorescein diacetate (DCFDA). Results: The ALA concentration corresponding to optimal cell uptake and PpIX production was determined to be 1mM, over a 24h incubation period. The photolysis of PpIX in cells irradiated with red LASER light was proven by the decrease of PpIX emission peak. The rate of ROS concentration increase (as determined with DCFDA) was diminished by adding vitamin C in the cellular suspension.
Key words: photodynamic therapy, 5-aminolaevulinic acid, protoporphyrin IX, red laser light, reactive oxygen species (ROS).
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