Abstract
The objective of this study was to investigate the effects of phototoxicity on retinal pigment epithelial (RPE) cells after zinc tetrasulfonated phthalocyanine bound bovine serum albumin (ZnPcS(4)-BSA) based photodynamic therapy (PDT). This study will provide a rational basis for the development of new clinical perspectives in future choroidal neovascularization (CNV) therapy. The 2-4 generation of subcultured RPE cells were used in this study. First, the cellular uptake of ZnPcS(4) and ZnPcS(4)-BSA was quantified by UV-spectra after confluent human RPE cells were exposed to various doses of ZnPcS(4) and ZnPcS(4)-BSA for 1-6 h. Second, RPE cells were divided into the following five groups: the control group, the group of photosensitizer without irradiation, the irradiated group without photosensitizer, PDT1 group (irradiated with 50 J/cm(2)), and the PDT2 group (irradiated with 100 J/cm(2)) that was exposed to 10 mg/mL ZnPcS(4)-BSA before they were irradiated. After ZnPcS(4)-BSA based PDT was performed, cell viability was evaluated by MTT, and reactive oxygen species (ROS) and mitochondrial membrane potential (Deltapsi(m)) were assessed by flow cytometry and confocal microscopy, respectively. The ratios of ZnPcS(4)-BSA content in the RPE cells were more than the intake ratios of ZnPcS(4) incubation at the same time point. It was also found that the optimal time point to irradiating the cell was 4 h after incubation. The cellular viability reduction ratios in the PDT1 group and PDT2 group RPE cell after ZnPcS(4)-BSA based PDT were 60.71% and 90.48%, respectively, which were higher than that of ZnPcS(4)-based PDT. PDT results in a significant increase in ROS. The mitochondrial membrane potentials of PDT1 group and PDT2 group were significantly decreased compared with the normal control group. The ratio of ZnPcS(4)-BSA content in RPE cells reached its peak after 4 h incubation. Intracellular ROS formation and the loss of Deltapsi(m) may play an important role in mediating the apoptotic processes of RPE cells induced by ZnPcS(4)-BSA based PDT. This study revealed that there is an important precondition to decrease the damage on RPE, that is, we should avoid irradiating RPE cells after the peak incubation time period.
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