29th Annual Meeting of the American Society of Photobiology

Downtown Marriot

Chicago, Il.

Comparison of apoptotic and necrotic response following 5-aminolaevulinic acid-based photodynamic therapy

Abstract-
The involvement of apoptosis has been implicated as an early event of cell death induced by photodynamic therapy (PDT) both in vitro and in vivo. However, the mode of cell death induced by PDT has been conflicted because of the cell lines used. While the biochemical mechanism of the enhanced protoporphyrin IX synthesis in the cancer cells following 5-aminolaevulinic acid (ALA) induction is well understood, little is known about the mechanism of ALA-PDT cytotoxicity. In this study, we investigated the ALA-PDT mediated cell death of human lung small cell carcinoma (Ms-1) and human promyelocytic leukemia (HL-60). First, we estimated the ALA-mediated photocytotoxicity. The two cell lines were equally sensitive to ALA-PDT when evaluated by MTT assay. Typical characteristics of apoptosis such as cell shrinkage and membrane blebbing were observed in the photosensitized HL-60 cells. On the other hand, such changes in morphology were not observed in Ms-1 cells. Next, we examined the effect of the caspase inhibitor (Z-asp-CH₂-DCB) on ALA-PDT-induced cell death. Inhibition of ALA-PDT-induced cell death was observed only in HL-60 cells. Furthermore, agarose gel electrophoresis revealed that only HL-60 cells undergo apoptosis, and the ladder pattern was diminished by addition of Z-asp-CH₂-DCB. Therefore, we have then examined proteolytic activation of caspase-3. When HL-60 cells were exposed to ALA-PDT, there was a dose-dependent loss of procaspase-3, indicative of the proteolytic processing of the proenzyme to the active enzyme subunits. These results suggest that ALA-PDT is capable of inducing apoptosis in a cell type-specific manner and caspase family was intimately involved in the ALA-PDT-induced apoptosis in HL-60 cells.

Keywords: photodynamic therapy, 5-aminolaevulinic acid, apoptosis, necrosis