Photodynamic Effect of Modulated and Continuous LASER Radiation on Cancerous Cells Cultivated In Vitro

Abstract

Photodynamic therapy (PDT) is a cancer therapy that uses nonionizing photons and a photosensitizer to treat solid tumors and surface malignancies. Experiments with cells in vitro have been useful in order to investigate cellular effects due to PDT. In this work, we study the photodynamic effect of modulated and continuous laser radiation on cancerous cells cultivated in vitro. Three experiments were performed for this study. For the first one, HeLa cells were exposed to δ-Aminolevulinic acid (ALA) at concentrations of 0, 10, 20, 40, 80 and 160 µg of ALA/ml of Dulbecco's modified Eagle's medium (DMEM) and were irradiated at total light dose of 80 J/cm2 by using continuous and modulated argon laser light. In the case of modulated radiation, at 1000 Hz, we observed more cellular death (20%) than those produced by continuous light (0%) when these cells were exposed at 20 µg of ALA/ml of DMEM. For other ALA doses cellular deaths were similar in both irradiation types. The death percentage in cancerous cell, after application of PDT with continuous and modulated irradiation, was determined by means of the neutral-red spectrophotometric assay. In the second experiment HeLa cells were exposed only at 20 µg of ALA/ml of DMEM and irradiated with argon laser at total light dose of 80 J/cm2 but using different light modulation frequencies: 100, 160, 600, 1250, 1500, 1850 Hz. Finally in the third experiment, where the HeLa cells were exposed in the same conditions of second experiment, the modulation frequencies used to irradiate the cell were: 10, 20, 40, 80 Hz. It was observed that light modulation frequency at 40 Hz produces maximum cellular death (36%). These results showed that modulated light used in PDT, for low ALA doses in HeLa cells, is more efficient than continuous light irradiation. Modulated irradiation in PDT could reduce the quantity of required ALA, and in consequence to reduce possible collateral damage, for application of this therapy.