Baldea Ioana, Olteanu Diana Elena, Bolfa Pompei, Tabaran Flaviu, Ion Rodica-Mariana, Filip Gabriela Adriana
University of Medicine and Pharmacy, Department of Physiology, Clinicilor 1, Cluj-Napoca, Romania.
University of Agricultural Sciences and Veterinary Medicine, Department of Pathology, Calea Manastur 3-5, 400372 Cluj-Napoca, Romania; Ross University School of Veterinary Medicine, Department of Biomedical Sciences, PO Box 334, Basseterre, St. Kitts, West Indies.
J Photochem Photobiol B. 2016 Aug;161:402-10. doi: 10.1016/j.jphotobiol.2016.06.012. Epub 2016 Jun 8.
Melanoma, a cancer derived from melanocytes is very difficult to treat, especially in advanced cases. There are several encouraging studies of the efficacy of photodynamic therapy (PDT) in melanoma. However, PDT has to overcome the main defense mechanisms like: defects in the apoptotic pathways, pigmentation, sequestration of the photosensitisers (PS) inside melanosomes and increased oxidative stress defense. Two meso-substituted porphyrins, meso-5,10,15,20-tetrakis (4-hydroxyphenyl) porphyrin (THOPP) and meso-5-(4-hydroxyphenyl)-10, 15, 20-tris (4-methoxyphenyl) porphyrin (THOMPP) were used as PS to investigate several mechanisms underlining the PDT anti-melanoma effects, on a lightly pigmented melanoma cell line (WM35), in vitro. γH2AX foci formation (a measure of DNA double strand brakes) was used for the assessment of DNA damage by means of immune-fluorescence and western blot. Cytoskeleton alterations were detected by phalloidin staining. Tyrosinase activity and melanin pigment were quantified by spectrophotometry, tyrosinase protein by western blot, total peroxidase activity by resorurfin reaction (Amplex Red). PDT induced high levels of DNA damage, cytoskeleton alterations and enhanced pigmentogenesis. THOPP mediated PDT was the most efficient. The melanogenesis stimulated by PDT was directly correlated to the PDT induced cellular damage and provided no protection against therapy. Thus, PDT induced melanogenesis combined with severe DNA damage was able to overcome the mechanisms of resistance and increased the efficiency of PDT in WM35 melanoma cells. These results are encouraging for a possible use of PDT, as an adjuvant therapy in lightly pigmented melanomas.
黑色素瘤是一种源自黑素细胞的癌症，很难治疗，尤其是在晚期病例中。有几项关于光动力疗法（PDT）治疗黑色素瘤疗效的研究令人鼓舞。然而，PDT必须克服主要的防御机制，如：凋亡途径缺陷、色素沉着、黑素小体内光敏剂（PS）的隔离以及氧化应激防御增强。两种中位取代卟啉，中位-5,10,15,20-四（4-羟苯基）卟啉（THOPP）和中位-5-（4-羟苯基）-10,15,20-三（4-甲氧基苯基）卟啉（THOMPP）被用作PS，以研究在体外对轻度色素沉着的黑色素瘤细胞系（WM35）进行PDT抗黑色素瘤作用的几种潜在机制。通过免疫荧光和蛋白质印迹法，利用γH2AX焦点形成（一种衡量DNA双链断裂的指标）来评估DNA损伤。通过鬼笔环肽染色检测细胞骨架的改变。通过分光光度法定量酪氨酸酶活性和黑色素，通过蛋白质印迹法定量酪氨酸酶蛋白，通过间苯二酚反应（Amplex Red）测定总过氧化物酶活性。PDT诱导高水平的DNA损伤、细胞骨架改变并增强色素生成。THOPP介导的PDT最有效。PDT刺激的黑色素生成与PDT诱导的细胞损伤直接相关，并且不能提供对治疗的保护作用。因此，PDT诱导的黑色素生成与严重的DNA损伤相结合能够克服耐药机制并提高PDT在WM35黑色素瘤细胞中的效率。这些结果对于将PDT作为轻度色素沉着黑色素瘤的辅助治疗方法的潜在应用来说是令人鼓舞的。
