Quantitative analysis for intracellular distribution of a photosensitizer using confocal laser scanning microscope
Confocal laser scanning microscope (CLSM) is capable of delivering a high axial resolution, and with this instrument even thin layers of cells can be imaged in good quality. Therefore, intracellular uptake and distribution properties of photosensitizer zinc coproporphyrin III tetrasodium salt (Zn CP-III) in human lung small cell carcinoma (Ms-1) were examined by using CLSM. In particular, the uptake of Zn CP-III in cytoplasm, plasma membrane, and nucleus was individually evaluated for the first time from fluorescence images obtained by CLSM. The results show that the Zn CP-III content in three cellular areas correlates with extracellular Zn CP-III concentration and time of incubation with Zn CP-III. Furthermore, it was found that the cytoplasmic fluorescence was approximately two times higher than that in the nucleus under all uptake conditions. In addition, cellular accumulation of Zn CP-III was compared with photodynamic cytotoxicity. The photocytotoxicity was to a great extent dependent on the uptake of the photosensitizer. The damaged site of Ms-1 cells induced by photodynamic therapy, was plasma membrane, However, the content of Zn CP-III accumulated in cytoplasm was the highest among the three areas, implying that, besides the direct damage on plasma membrane, an oxidative damage to cellular component arose from the cytoplasmic Zn CP-III may also play an important role in photocytoxicity. The quantitative information obtained in this study will be useful for further investigation of the photocytotoxicity as well as the uptake mechanism of photosensitizer.
photosensitizer photodynamic therapy confocal laser scanning microscopy image analysis