PHOTODYNAMIC EFFECTS ON HUMAN AND CHICKEN ERYTHROCYTES STUDIED WITH MICROIRRADIATION AND CONFOCAL LASER-SCANNING MICROSCOPY

Background and Objective: Photodynamic therapy (PDT) of cancers is ass ociated with the destruction of the microvasculature supplying the tum or. The study elucidates the role of red blood cells in PDT-induced va scular injury. Study Design/Materials and Methods: Intracellular accum ulation of several photosensitizers in human (non-nucleated) and chick en (nucleated) erythrocytes, as well as photodynamic induced hemolysis were studied using 488 nn laser microirradiation (15 mu W) and confoc al laser scanning fluorescence microscopy. Results: Cells incubated wi th anionic hydrophilic compounds TPPS4 and Pd-TPPS4 exhibited no fluor escence before irradiation, but developed strong and sustained fluores cence in the cellular and nuclear membranes following photoinduced mem brane damage. In contrast, microirradiation of Photofrin-incubated ery throcytes showed instantaneous fluorescence which decreased due to pho todegradation. For the cationic hydrophilic dye methylene blue, signif icant fluorescence was detected only in the nucleus. Following ALA inc ubation, large intercellular differences were observed in fluorescence in the red spectral region. Photofrin induced the most efficient hemo lysis. Higher radiant exposures were required for lysis of nucleated r ather than of non-nucleated red blood cells, except in the case of met hylene blue. Conclusion: Laser microbeams were used, for the first tim e, to study photodynamic cell damage. Erythrocytes were shown to be pr imary targets in PDT. Damage to red blood cells could be responsible f or hemostasis in the vascular bed of a tumor, which was reported by ma ny groups. (C) 1996 Wiley-Liss, Inc.