Photodynamic activity of a new sensitizer derived from porphyrin-C60 dyad and its biological consequences in a human carcinoma cell line
Photodynamic activity of a new sensitizer derived from porphyrin-C60 dyad and its biological consequences in a human carcinoma cell line
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DOI：
10.1016/j.biocel.2006.05.019
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发表时间：
2006-01-01
影响因子：
4
通讯作者：
Rivarola, Viviana
中科院分区：
文献类型：
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作者：
Alvarez, M. Gabriela;Prucca, Cesar;Rivarola, Viviana
以人喉癌Hep-2细胞株为研究对象，评价了一种卟啉-C-60(P-C-60)二聚体的光杀伤活性。这项研究首次评价了一种具有高能力形成光诱导电荷分离状态的二聚体，作为光动力疗法(PDT)灭活细胞的试剂。细胞处理采用1muM-P-C-60脂质体包埋。1 mU M P-C-60浓度和长时间孵育(24 H)均未观察到暗细胞毒性。在不同的孵育时间内，研究增敏剂对Hep-2细胞的摄取。在此条件下，细胞孵育4h后，细胞存活率为1.5nmol/10(6)个细胞，24小时后细胞存活率基本保持不变。P-C-60具有较强的细胞毒作用，54J/cm~(2)照射后细胞失活达80%以上。此外，该二聚体在Ar气氛下仍保持较高的光活性。因此，根据敏化剂所处的微环境，该化合物在低氧浓度下可以通过O-1(2)介导的光反应过程或自由基机制产生生物光损伤。通过Hoechst-33258、甲苯胺蓝染色、TUNEL和DNA片段分析了细胞死亡的机制。用P-C-60处理细胞24 h，54J/cm(2)照射后，细胞出现大量的凋亡细胞(58%)。用Hoechst-33258荧光显微镜观察细胞在低氧浓度下的形态变化。在这种厌氧条件下，细胞的死亡以坏死性死亡为主。空气照射条件下的细胞凋亡率明显高于氩气照射条件下的细胞凋亡率。用caspase-3活性检测试剂盒检测caspase-3活性，以确定细胞的凋亡途径。结果表明，P-C-60通过caspase-3依赖途径诱导细胞凋亡。这些结果表明，分子二聚体可以形成光诱导电荷分离态，是一种很有前途的光疗剂模型，在光动力灭活细胞方面具有潜在的应用前景。(C)2006爱思唯尔有限公司。保留所有权利。
The photokilling activity of a porphyrin-C-60 (P-C-60) dyad was evaluated on a Hep-2 human larynx-carcinoma cell line. This study represents the first evaluation of a dyad, with high capacity to form a photoinduced charge-separated state, to act as agent to inactivate cells by photodynamic therapy (PDT). Cell treatment was carried out with 1 mu M P-C-60 incorporated into liposomal vesicles. No dark cytotoxicity was observed using 1 mu M P-C-60 concentration and during long incubation time (24 h). The uptake of sensitizer into Hep-2 was studied at different times of incubation. Under these conditions, a value of 1.5 nmol/10(6) cells was found after 4 h of incubation showing practically no change even after 24 h. The cell survival after irradiation of the cells with visible light was dependent upon light exposure level. A high photocytotoxic effect was observed for P-C-60, which inactivated 80% of the cells after 54 J/cm(2) of irradiation. Moreover, the dyad kept a high photoactivity even under argon atmosphere. Thus, depending on the microenviroment where the sensitizer is localized, this compound could produce a biological photodamage through either a O-1(2)-mediated photoreaction process or a free radical mechanism under low oxygen concentration.The mechanism of cell death was analyzed by Hoechst-33258, toluidine blue staining, TUNEL and DNA fragmentation. Cell cultures treated for 24 h with P-C-60 and irradiated with a dose of 54 J/cm(2) showed a great amount of apoptotic cells (58%). Moreover, changes in cell morphology were analyzed using fluorescence microscopy with Hoechst-33258 under low oxygen concentration. Under this anaerobic condition, necrotic cellular death predominated on apoptotic pathway. There were more apoptotic cells under air irradiation condition than under argon irradiation condition. To determine the apoptotic pathway, caspase-3 activation was studied by caspase-3 activity detection kits. The last results showed that P-C-60 induced apoptosis by caspase-3-dependent pathway. These results indicated that molecular dyad, which can form a photoinduced charge-separated state, is a promising model for phototherapeutic agents and they have potential application in cell inactivation by PDT. (c) 2006 Elsevier Ltd. All rights reserved.