Article
5-ALA based photodynamic therapy (PDT) in glioblastoma-derived stem cell-enriched cultures
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Published: | September 16, 2010 |
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Objective: Considerable attention is being focussed on basic and translational research concerning glioblastoma stem cells, their tumorigenic potential and their resistance to radio- and chemotherapy. 5-ALA based photodynamic therapy (PDT) induces cell death in the C6 glioma spheroid model and appears to improve clinical outcome in patients suffering from recurrent glioblastoma by causing long-sustaining responses. In order to study the acute effects of ALA/PDT on multipotent glioblastoma cell lines, we used a three-dimensional cell culture system.
Methods: Glioblastoma-derived stem cell-enriched cultures were treated with ALA/PDT. Human glioma spheroids (U 373) served as controls. Cell spheroids were incubated for 4 h in 0/4/12,5/18 µg/ml ALA in 5% CO2 in room air with subsequent irradiation using a diode laser (lambda = 635 nm, 40 mW/cm2, total fluence 25 J/cm2). Control groups were "laser only", "ALA only", and "no drug no light".The stem cell lines were analyzed for expression profiles like CD133 and its intracellular concentration after ALA-incubation by fluorescence activated cell sorting (FACS). Cell death was analyzed by colorimetric assay (WST-1 based) for the nonradioactive quantification of cell proliferation, cell viability and cytotoxicity.
Results: Subsequent ALA/PDT after incubation in 5% CO2 provided incomplete cell death in tumor spheroids in relation to “no drug, no light” controls: 0µg/ml ALA: 99,54%, 4 µg/ml ALA: 122,02%, 12,5 µg/ml ALA: 42,45% and 18 µg/ml ALA: 21,00%. In incompletely damaged spheroids viable cells were restricted to spheroid centers. The rate of cell death in all control groups was negligible. The cytotoxicity of the stem cells after ALA/PDT was equivalent to those results of the U373 glioma spheroids. Expression of CD133 was confirmed in all stem cell lines with typical full spheroid growth pattern.
Conclusions: PDT of experimental glioblastoma stem cell lines results in significant cell death after irradiation. This might be an explanation for observed long-sustaining PDT effects in recurrent glioblastomas in patients.