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Photodynamic effects on human and chicken erythrocytes studied with microirradiation and confocal laser scanning
1Beckman Laser Institute and Medical Clinic, UCI, Irvine, California 92715, USA.
Background And Objective:
Photodynamic therapy (PDT) of cancers is associated with the destruction of the microvasculature supplying the tumor. The study elucidates the role of red blood cells in PDT-induced vascular injury.
Study Design/Materials And Methods:
Intracellular accumulation of several photosensitizers in human (non-nucleated) and chicken (nucleated) erythrocytes, as well as photodynamic induced hemolysis were studied using 488 nm laser microirradiation (15 microW) and confocal laser scanning fluorescence microscopy.
Results:
Cells incubated with anionic hydrophilic compounds TPPS4 and Pd-TPPS4 exhibited no fluorescence before irradiation, but developed strong and sustained fluorescence in the cellular and nuclear membranes following photoinduced membrane damage. In contrast, microirradiation of Photofrin-incubated erythrocytes showed instantaneous fluorescence which decreased due to photodegradation. For the cationic hydrophilic dye methylene blue, significant fluorescence was detected only in the nucleus. Following ALA incubation, large intercellular differences were observed in fluorescence in the red spectral region. Photofrin induced the most efficient hemolysis. Higher radiant exposures were required for lysis of nucleated rather than of non-nucleated red blood cells, except in the case of methylene blue.
Conclusion:
Laser microbeams were used, for the first time, to study photodynamic cell damage. Erythrocytes were shown to be primary targets in PDT. Damage to red blood cells could be responsible for hemostasis in the vascular bed of a tumor, which was reported by many groups.