Li Wen-Tyng, Tsao Hsin-Wei, Chen Ying-Ying, Cheng Shih-Wei, Hsu Yih-Chih
Department of Biomedical Engineering, Chung-Yuan Christian University, Chung-Li, Taiwan, R.O.C.
Photochem Photobiol Sci. 2007 Dec;6(12):1341-8. doi: 10.1039/b704539e. Epub 2007 Oct 16.
Photodynamic therapy (PDT) is an alternative anticancer treatment in which direct tumor-cell killing results from selective accumulation of photosensitizers in the tumor sites and phototoxicity occurs when light-activated photosensitizers transfer the energy to oxygen nearby to produce singlet oxygen. The objective of this study was to investigate the effects of PDT using chlorophyll derivatives such as pheophytin a (phe a), pheophytin b (phe b), pheophorbide a (pho a) and pheophorbide b (pho b) as the photosensitizers, and the 660 nm light-emitting diodes (LEDs) irradiation on human hepatocellular carcinoma cells (HuH-7). The drug concentration-dependent inhibition of HuH-7 cell viability was studied under LEDs irradiation (10 mW cm(-2)) at radiant exposure of 5.1 and 10.2 J cm(-2) by MTT assay. Significant inhibition of the survival of HuH-7 cells (<10%) was observed when an irradiation dose of 10.2 J cm(-2) combined with the concentration of 0.5 microg ml(-1) of phe a, 0.125 microg ml(-1) of pho a, 0.25 microg ml(-1) of phe b, and 0.125 microg ml(-1) of pho b were applied. The results from Annexin V-propidium iodide staining revealed that phe a, phe b, pho a and pho b could induce cell death in HuH-7 cells predominantly via a necrotic process. The results from immunoblot analyses exhibited that chlorophyll derivative-mediated PDT initiated cytochrome c release, caspase-9 and caspase-3 activation, followed by poly ADP-ribose polymerase (PARP) cleavage. Thus, apoptosis also occurred in HuH-7 cells after PDT treatment, and the execution of the apoptotic process may be initiated from the loss of mitochondrial function. Our findings demonstrate that both apoptosis and necrosis can be induced in HuH-7 cells after PDT using phe a, phe b, pho a and pho b and LEDs.
光动力疗法（PDT）是一种替代性抗癌治疗方法，其中肿瘤细胞的直接杀伤源于光敏剂在肿瘤部位的选择性积聚，当光激活的光敏剂将能量转移至附近的氧气以产生单线态氧时，就会发生光毒性。本研究的目的是调查使用叶绿素衍生物如脱镁叶绿酸a（phe a）、脱镁叶绿酸b（phe b）、脱镁叶绿甲酯酸a（pho a）和脱镁叶绿甲酯酸b（pho b）作为光敏剂的光动力疗法以及660纳米发光二极管（LED）照射对人肝癌细胞（HuH-7）的影响。通过MTT法在5.1和10.2 J cm⁻²的辐射暴露下，在LED照射（10 mW cm⁻²）条件下研究了HuH-7细胞活力的药物浓度依赖性抑制作用。当应用10.2 J cm⁻²的照射剂量并结合0.5 μg ml⁻¹的phe a、0.125 μg ml⁻¹的pho a、0.25 μg ml⁻¹的phe b和0.125 μg ml⁻¹的pho b浓度时，观察到HuH-7细胞存活率受到显著抑制（<10%）。膜联蛋白V-碘化丙啶染色结果显示，phe a、phe b、pho a和pho b主要通过坏死过程诱导HuH-7细胞死亡。免疫印迹分析结果表明，叶绿素衍生物介导的光动力疗法引发细胞色素c释放、半胱天冬酶-9和半胱天冬酶-3激活，随后是聚ADP-核糖聚合酶（PARP）裂解。因此，光动力疗法治疗后HuH-7细胞中也发生了凋亡，并且凋亡过程的执行可能始于线粒体功能的丧失。我们的研究结果表明，使用phe a、phe b、pho a、pho b和LED进行光动力疗法后，HuH-7细胞中可同时诱导凋亡和坏死。
