Programa de Pós-Graduação Interunidades Bioengenharia EESC/FMRP/IQSC, Universidade de São Paulo, São Carlos, SP, Brazil.
Departamento de Diagnóstico e Cirurgia, Universidade Estadual Paulista Júlio de Mesquita Filho, Araraquara, SP, Brazil.
Photodiagnosis Photodyn Ther. 2017 Sep;19:355-362. doi: 10.1016/j.pdpdt.2017.07.003. Epub 2017 Jul 14.
Photodynamic therapy (PDT) has been established in several countries as an alternative therapy for the treatment of various malignancies. This therapy involves the incorporation of a photosensitizer (PS) that is activated by visible light and form reactive oxygen species leading to target cell death by apoptosis or necrosis. Previously, our group has demonstrated that CHL-T (semi-synthesized from chlorophyll a and containing a linked solubilizing group TRISMA) presented a pronounced potential to induce death in HeLa cell line after PDT. In the present study, besides confirm the high cytotoxicity in another cell line, we have further investigated the cell death mechanisms caused by CHL-T as a photosensitizer in laryngeal carcinoma cells.
Cells were exposed to different concentrations of three photosensitizers, namely, hypericin (HY), unmodified chlorin (CHL) and a synthesized amphiphilic chlorin derivative (CHL-T). PSs accumulation and localization were accessed by fluorescence assays. Photosensitization was induced at 6Jcm using red LEDs (630±10nm). Viability was assessed by mitochondrial function (MTT); whereas apoptosis/necrosis was evaluated by fluorescence microscopy and flow cytometry. Expression of pro-apoptotic p53 protein was studied by Western blot.
All PS showed similar localization profile in the HEp-2 cells. The use of CHL-T increased the percentage of apoptotic cells and also p53 expression in comparison with the use of HY and CHL as photosensitizers. This study shows a significant effect of CHLT associated with red light (630±10nm and 18mWcm) irradiation on a cancer cell line, indicating the potential of this amphiphilic chlorin in enhancing the therapeutic effectiveness of Photodynamic Therapy (PDT).
光动力疗法(PDT)已在多个国家被确立为治疗各种恶性肿瘤的替代疗法。该疗法涉及到光敏剂(PS)的掺入,该光敏剂被可见光激活,并形成活性氧物种,导致靶细胞通过细胞凋亡或坏死死亡。此前,我们的研究小组已经证明,CHL-T(由叶绿素 a 半合成并含有连接的增溶基团 TRISMA)在 PDT 后对 HeLa 细胞系表现出明显的致死潜力。在本研究中,除了在另一种细胞系中确认高细胞毒性外,我们还进一步研究了 CHL-T 作为光敏剂在喉癌细胞中引起细胞死亡的机制。
将细胞暴露于三种光敏剂(即金丝桃素(HY)、未修饰的叶绿素(CHL)和合成的两亲性叶绿素衍生物(CHL-T))的不同浓度下。通过荧光测定法评估 PS 的积累和定位。在 630±10nm 的红色 LED 下用 6J/cm 的光进行光致敏。通过线粒体功能(MTT)评估细胞活力;而通过荧光显微镜和流式细胞术评估凋亡/坏死。通过 Western blot 研究促凋亡 p53 蛋白的表达。
所有 PS 在 HEp-2 细胞中均显示出相似的定位模式。与使用 HY 和 CHL 作为光敏剂相比,使用 CHL-T 增加了凋亡细胞的百分比和 p53 表达。这项研究表明,CHLT 与红光(630±10nm 和 18mW/cm)照射相结合对癌细胞系有显著影响,表明这种两亲性叶绿素在增强光动力疗法(PDT)的治疗效果方面具有潜力。