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Induction of apoptotic cell death by photodynamic therapy in human keratinocytes
A Ketabchi1, A MacRobert, P M Speight
1Department of Oral Pathology, Eastman Dental Institute, London.
Abstract:
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.
Insights
Photodynamic therapy (PDT) with aluminium disulphonated phthalocyanine (AlS2Pc) effectively induces apoptosis, or programmed cell death, in skin and oral cancer cells. This research clarifies PDT
Area of Science:
- Oncology
- Photochemistry
- Cell Biology
Background:
- Photodynamic therapy (PDT) is used clinically for skin and oral cancers, but its precise mechanism of action requires further scientific elucidation.
- Existing evidence suggests epithelial cell death in PDT may result from vascular damage or direct cell killing via apoptosis-like pathways.
Purpose of the Study:
- To investigate the mechanism of epithelial cell death induced by photodynamic therapy (PDT) in vitro.
- To determine if PDT with aluminium disulphonated phthalocyanine (AlS2Pc) promotes apoptosis in human epidermal keratinocytes and oral squamous cell carcinoma cells.
Main Methods:
- Human epidermal keratinocytes (UP) and oral squamous cell carcinoma cells (H376) were treated with AlS2Pc and 675 nm red laser light.
- Cell viability was assessed using an MTS assay.
- Apoptosis was evaluated through morphological analysis (haematoxylin and eosin staining), ultrastructural examination, and in situ end-labeling of DNA fragments.
Main Results:
- PDT treatment significantly reduced cell viability in both UP and H376 cell lines (p < 0.01).
- A significant increase in the number of apoptotic cells was observed in PDT-treated cultures compared to controls.
- Ultrastructural analysis and DNA fragmentation assays confirmed apoptosis induction.
Conclusions:
- Photodynamic therapy utilizing AlS2Pc as a photosensitizer effectively induces apoptotic cell death in human epidermal keratinocytes and oral squamous cell carcinoma cells in vitro.
- These findings suggest that direct epithelial cell killing through apoptosis may be a significant contributor to tumor necrosis observed in vivo following PDT.