Berrahmoune Saoussen, Fotinos Nicolas, Bezdetnaya Lina, Lange Norbert, Guedenet Jean Claude, Guillemin François, D'Hallewin Marie Ange
Photobiology in Cancerology, Centre Alexis Vautrin, CRAN Nancy University, CNRS, Avenue de Bourgogne 54511, and Department of pathology and cytology, University Hospital Nancy, Vandoeuvre les Nancy Cedex, France.
Photochem Photobiol Sci. 2008 Sep;7(9):1018-24. doi: 10.1039/b804921a. Epub 2008 Jul 23.
The hexylester of 5-aminolevulinic acid (HAL) is a very efficient precursor of the photosensitizer protoporphyrin IX (PpIX) for photodynamic therapy (PDT). Our previous study, performed in rat orthotopic bladder tumors, indicated an opposite effect of HAL/PpIX-PDT according to HAL concentration. The present study investigated possible reasons for this differential effect considering the impact of extracted amounts of PpIX in normal and tumor bearing bladders along with PpIX distribution in distinctive histopathological layers. High performance liquid chromatography (HPLC) analysis of tumor and normal bladder tissues after 8 mM and 16 mM HAL instillation showed that PpIX was the main porphyrin species. The PpIX production in tumor bladders instilled with 8 mM HAL was significantly higher than after 16 mM HAL. Fluorescence confocal microscopy demonstrated a punctuate bright fluorescence pattern in tumor zones of bladders instilled with 8 mM HAL, whereas a more diffuse cytoplasmatic fluorescence distribution was observed after 16 mM HAL instillation. Immunofluorescence staining together with transmission electron microscopy showed severe mitochondrial damage in tumor zones of bladders treated with 8 mM HAL/PpIX PDT, with intact mitochondria in tumor zones of bladders treated with 16 mM HAL/PpIX PDT. We conclude that the differential response to HAL/PpIX PDT in function of HAL concentrations could be attributed to diminished PpIX synthesis and differential intracellular localisation of PpIX. Mitochondria were shown to be the critical photodamaged sites of HAL/PpIX PDT and as such tissue sensitivity to treatment can be estimated through investigation of intracellular PpIX distribution.
5-氨基乙酰丙酸己酯（HAL）是用于光动力疗法（PDT）的光敏剂原卟啉IX（PpIX）的一种非常有效的前体。我们之前在大鼠原位膀胱肿瘤中进行的研究表明，根据HAL浓度不同，HAL/PpIX-PDT会产生相反的效果。本研究考虑了正常膀胱和荷瘤膀胱中PpIX提取量的影响以及PpIX在不同组织病理学层面的分布，探讨了这种差异效应的可能原因。对分别灌注8 mM和16 mM HAL后的肿瘤和正常膀胱组织进行高效液相色谱（HPLC）分析表明，PpIX是主要的卟啉种类。灌注8 mM HAL的肿瘤膀胱中PpIX的生成量显著高于灌注16 mM HAL后。荧光共聚焦显微镜显示，灌注8 mM HAL的膀胱肿瘤区域呈现点状明亮荧光模式，而灌注16 mM HAL后观察到更弥漫的细胞质荧光分布。免疫荧光染色结合透射电子显微镜显示，用8 mM HAL/PpIX PDT处理的膀胱肿瘤区域线粒体严重受损，而用16 mM HAL/PpIX PDT处理的膀胱肿瘤区域线粒体完整。我们得出结论，HAL/PpIX PDT对HAL浓度的不同反应可能归因于PpIX合成减少和PpIX在细胞内的定位差异。线粒体被证明是HAL/PpIX PDT的关键光损伤部位，因此可以通过研究细胞内PpIX分布来评估组织对治疗的敏感性。
