Induction of apoptotic cell death by photodynamic therapy in human keratinocytes.

The use of photodynamic therapy (PDT) for the treatment of skin and oral cancer has been the subject of several clinical studies but there has been little scientific evaluation of its mechanism of action. Evidence to date suggests that whilst epithelial cell death may be secondary to vascular damage, direct cell killing may occur and may involve an apoptosis-like mechanism. To investigate the mechanism of epithelial cell death following PDT, two cell lines, human epidermal keratinocytes (UP) and oral squamous cell carcinoma-derived cells (H376) were subjected to PDT with aluminium disulphonated phthalocyanine (AlS2Pc) as the photosensitizer and red laser light at 675 nm. Control groups received red laser light, photosensitizer or neither. The effects of PDT were assessed using an MTS cell-proliferation assay, which showed a significant reduction in viability (p < 0.01) for PDT-treated cells compared to controls. For morphological analysis, cells were stained with haemotoxylin and eosin and the numbers showing typical apoptotic features counted. The treated cultures showed significantly increased numbers of apoptotic cells. Moreover, the H376 control cultures showed a baseline level of apoptosis of approx. 15%. Apoptosis was confirmed by ultrastructural analysis and by in situ end-labeling of DNA fragments. The results show that PDT using AlS2Pc as a photosensitizer promotes apoptotic cell death in UP and H376 cells in vitro and suggest that direct killing of epithelial cells may contribute to tumour necrosis in vivo.