Photodynamic Therapy in Hep-2 Cell

ÁLVAREZ, M. G; RIVAROLA, V.; FUKUDA, H.; BATLLE, A.

Shanghai, China

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

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 and 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 mM P-C (60 ), 5 mM 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 P-C(60) 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.