CELLULAR & SUBCELLULAR MECHANISMS OF PHOTODYNAMIC THERAPY

在这个项目中，我们正在研究的基本机制， 光动力疗法（PDT）对肿瘤微结构的作用。 具体而言，亚细胞定位和亚细胞 PDT药物的光毒性正在研究中。 首先，CPAE 内皮细胞和PTK2上皮细胞的研究，通过可视化， 当细胞暴露于 5-氨基乙酰丙酸（ALA）。 两种细胞类型都显示出大量的 在核周细胞质中的荧光， 光敏剂选择性地定位在线粒体中。 到 研究ALA的亚细胞光毒性， 用630 nm激光微束照射细胞区域。 结果表明，核周细胞的数量最多，其次是核周细胞 细胞质是细胞中最敏感的区域。 特别 有趣的是发现细胞核是最敏感的 区域，因为1）在细胞核中没有观察到荧光，2） PDT损伤普遍接受机制涉及单线态氧 细胞质元件的靶向。 根据这些发现，我们将 进一步研究亚细胞定位和光毒性 ALA和其他PDT药物使用双光子激发。 PDT的激活 药物通过多光子吸收提供了更大的选择性， 单光子吸收 此外，双光子荧光将 允许改善细胞内药物分布的可视化， 尤其是沿着光轴。

In this project we are investigating the fundamental mechanisms of photodynamic therapy (PDT) action on tumor microstructure. Specifically, the subcellular localization and subcellular phototoxicity of PDT drugs are being investigated. Initially, CPAE endothelial and PTK2 epithelial cells were studied by visualizing the subcellular organelle fluorescence when the cells were exposed to 5-aminolaevulinic acid (ALA). Both cell types showed a large amount of fluorescence in the perinuclear cytoplasm, where the photosensitizer selectively localized in the mitochondria. To investigate the subcellular phototoxicity of ALA, the subcellular regions of the cells were irradiated with a 630 nm laser microbeam. The results demonstrated that the nucleus followed by the perinuclear cytoplasm are the most sensitive areas of the cell. Particularly intriguing was the finding that the nucleus was the most sensitive region since 1) no fluorescence was observed in the nucleus and 2) th e generally accepted mechanism for PDT damage involve singlet oxygen targeting of cytoplasmic elements. Based upon these findings we will further investigate the subcellular localization and phototoxicity of ALA and other PDT drugs using 2-photon excitation. Activation of PDT drugs through multiphoton absorption provides greater selectivity than single photon absorption. in addition, the 2-photon fluorescence will allow improved visualization of drug distribution within the cell, especially along the optical axis.