Acridine orange-mediated photodamage to cultured cells

Photosensitization mediated by the lysosomotropic, weakly basic dye acridine orange (AO) was studied on cultured J-774 cells. The phototoxicity was found to be potentiated by elevated oxygen tension and reduced at low oxygen tension. Moreover, cell cultures pre-exposed to the singlet oxygen scavenger sodium azide showed pronounced protection against the loss of viability induced by AO and blue light. AO-mediated photosensitization was neither increased by pre-exposure of cell cultures to ferric chloride or the catalase-inhibitor aminotriazole nor decreased by exposure to deferoxamine. These observations suggest that type II (singlet oxygen-mediated) reactions predominate over type I reactions (radical-mediated). A rapid and pronounced decrease in lysosomal cathepsin L activity (up to 60%) was observed after an initial 10 min irradiation, indicating the lysosomal compartment to be an early target. This irradiation time did not, however, result in any substantial loss of viability. Levels of cytosolic lactate dehydrogenase were unaffected even after 30 min irradiation, indicating that neither cytosol nor plasma membrane is a primary target of the AO-mediated photodamage. Glutathione depletion by pre-exposure to buthionine-S,R-sulfoximine (BSO) much enhanced the sensitivity of J-774 cells to AO-mediated photosensitization, indicating a protective role for thiol-containing compounds against AO-mediated photodamage.