荧光探针在光动力疗法亚细胞损伤位点研究中的应用

目的：应用荧光探针在光动力疗法研究中检测亚细胞损伤位点。方法：传代培养鼠肺毛细血管内皮细胞，将血卟啉单甲醚(HMME)与内皮细胞共同孵育24小时后，加入线粒体探针Bhodamine-123、内质网探针DioC6(3)和溶酶体探针Lucifer yellow分别对细胞器染色。首先采用激光共聚焦显微镜对光敏剂进行亚细胞定位。应用荧光显微镜汞灯照射激发光敏剂的光动力效应，加入ROS探针H2DCF-DA检测产生的单线态氧。分别在激发前后采集Pdloclamine-123、Lucifer yellow和DioC6(3)的荧光图像。结果：线粒体探针Phodamine-123的荧光图像在光动力损伤前后差异显著，原有形态特点发生明显改变；Phodamine-123在光动力损伤后再分布于细胞核区。结论：血卟啉单甲醚介导的光动力效应导致亚细胞水平多位点损伤，线粒体和核膜可能是PDT敏感位点；荧光探针标记检测光动力损伤亚细胞位点方法简便可靠。

Applving Fluorescence Probes in Studying Subcellular Photodamage Sites

Objective: To study the subcellular photodamage sites of hematoprophyrin monomethyl ether (HMME) PDT in murine lung endothelial cells applying fluorescence probes.Methods: Murine lung endothelial cells were subcultured and incubated with HMME for 24 hours. Then the cells were stained with Rhodamine 123 for mitochondrion? Lucifer yellow for lysosome?DioC 6(3) for ER. Laser Scanning Confocal Microscope was applied to study subcellular localization of HMME. H2DCF-DA was used to label singlet oxygen after irradiation by mercury lamp. Fluorescence images of Rhodamine 123? Lucifer yellow and DioC 6(3) were recorded respectively before and after irradiation. Results: Difference of Rhodamine 123's Fluorescence images is significant before and after photodamage.The formal character of them has changed obviously. Rhodamine 123 redistributed in nucleu after irradiation. nucleu may be susceptible targets;Studying subcelluar photodamage sites by labeling with Fluorescence probes is simple and reliable.