Light-induced apoptosis involves a defined sequence of cytoplasmic and nuclear calcium release in AlPcS4-photosensitized rat bladder RR 1022 epithelial cells

Oxidative stress induced by light activation of photosensitizers is regarde d to have a role in triggering cell death pathways during photodynamic ther apy (PDT). Reactive oxygen species have been proposed to act as signal tran sduction molecules activating downstream reactions that lead to apoptosis, Mainly debated is the cooperating role of other signaling systems like calc ium or pH. The present work contributes to this discussion by studying PDT effects in cell cultures of rat bladder epithelial cells for the hydrophili c tetrasulfonated aluminum phthalocyanine (AlPcS4), Cells were coincubated with the photosensitizer and the calcium-sensitive probe Fluo-3, The light- induced reactions were analyzed with a confocal laser scanning microscope. The dynamics of the process during light activation was observed with subce llular resolution. A transient calcium elevation during the irradiation pro cess was detected, especially in the cell's nuclei, followed by a more sust ained increase. The evaluation of the energy-dose-dependent phototoxicity a fter an incubation time with the photosensitizer of I and 24 h, showed enha nced phototoxicity when the drug was present for 24 h, Surprisingly, stimul ation of cell proliferation was observed at very low light doses (at 0.2 J/ cm(2)) when the drug was incubated for 24 h (cell viability 160%), Inductio n of apoptosis could be observed after irradiation with fluences between 1 and 3 J/cm(2). Apoptotic cells were identified with fluorescein isothiocyan ate-labeled Annexin V, which binds to phosphatidylserine after its transloc ation to the outer plasma membrane. In the presence of the antioxidant pyrr olidinedithiocarbamate the transient calcium elevation was totally inhibite d, as was the subsequent translocation of PS, In contrast, N-acetyl-L-cyste ine did not suppress the transient calcium increase. Our data might be cons istent with calcium regulated processes during AlFcS(4)-PDT and the involve ment of oxygen radicals.