The type-II photosensitization process is mediated by the formation of singlet oxygen (O2[1deltag]). The short lifetime of this species dictates that chemical reactions with biological substrates can only occur when O2(1deltag) is in very close proximity to the photosensitizer itself. In this study, deuteroporphyrin, a type-II, membrane-localized photosensitizer, was used to generate O2(1deltag) in human lymphoblast WTK-1 cells, and the range of influence was determined by a variety of biological assays. Surprisingly, the initial membrane-confined events were shown, by comet assay, to induce DNA damage in these cells. DNA damage was inhibited both by membrane-localized (alpha-tocopherol acetate) and by cytoplasmic (trolox) free radical scavengers. Comet formation also was inhibited by treatment at low temperature. DNA fragmentation was not influenced by treatment with the pan-caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone, showing that apoptosis was not responsible for fragmentation. Taken together, these results show that primary photosensitization reactions involving O2(1deltag), even when tightly confined in extranuclear locations, leads to the production of secondary reactive oxygen species, probably as a result of lipid peroxidation, that can act at greater distances from the photosensitizer itself. These experiments were carried out under conditions where cell survival was significant and raise questions regarding DNA damage and mutagenesis pathways, even when extranuclear O2(1deltag)-generating compounds are used.

No abstract

The fluoroquinolone antibiotics are widely used despite their strong phototoxicity under solar UV irradiation. Although they are known as good photodynamic photosensitizers, other factors than production of activated oxygen species may play a role in the effectiveness of the phototoxic effect. Subcellular localization is one of the important parameters that may determine this strength. Using microspectrofluorometry, it is shown that norfloxacin, ofloxacin, lomefloxacin, ciproflaxin and BAYy3118 are readily incorporated into lysosomes of HS68 human skin fibroblasts although weak staining of the whole cytoplasm also occurs especially with norfloxacin. Consistent with their photoinstability in solutions, the fluoroquinolones under study are readily photobleached by UVA in the HS68 fibroblasts. The BAYy3118 derivative that has the fastest bleaching rate also shows the strongest phototoxicity toward HS68 fibroblasts. Photosensitization with these fluoroquinolones induces lysosomal membrane damage as shown by the increased rate of leakage of the lysosomal probe lucifer yellow as compared to that observed with untreated cells.