Photodynamic Therapy in Hep-2 cells

ALVAREZ, GABRIELA; RIVAROLA, VIVIANA; FUKUDA, HAYDéE; BATLLE, ALCIRA

Shanghai, China

Congreso; 11th World Congress of the International Photodynamic Association - IPA 2007; 2007

International Photodynamic Association

Introduction: All living cells form Haem from Aminolevulinic acid (ALA). After exogenous administration, ALA is converted into the photosensitiser (PS) Protoporphyrin IX (PPIX).  PPIX is accumulated in malignant cells. After exposure to light, activated PPIX generates reactive oxygen species which conduce to cell death, the so-called Photodynamic Therapy (PDT).In this work we have evaluated the photokilling activity of three PSs on the Hep-2 cells. Materials & Methods: The Hep-2 human larynx carcinoma cell line and the following PSs were used: molecular dyad porphyrin C-(60) (P-C (60)); the monocationic porphyrin derivative 5-(4-trimethyl ammoniumphenyl)-10, 15, 20-tris (2, 4, 6-trimethoxyphenyl) porphyrin (CP) and PPIX synthesized in situ from ALA. The mechanism of cell death was analyzed by Hoechst -33258 and toluidine blue staining, TUNEL assay and DNA fragmentation. Results: No dark cytotoxicity was observed with 1 mìM PC (60), 5 ìm CP or 1 mm ALA along the 24 h incubation. Cell survival after irradiation with visible light was dependent on incubation time. Cells treated 24 h with P-C (60) and irradiated with 54 J/cm2 showed 58 % of apoptosis. Instead, under these conditions, CP induced a 58 % of necrosis. PPIX accumulated after incubation with 1 mM ALA for 45 h induced 40 % of apoptosis. Upon irradiation in anaerobiosis of cells incubated with P-C (60), necrosis predominated over apoptosis.   Microscopy under violet-blue light revealed that fluorescence of CP was localized in mitochondria, while ALA-induced PPIX was localized in lysosomes. Conclusion: Photodamage in PDT is dependent on the PS, light dose and the atmosphere conditions.