Abstract：
Malignant melanoma is the most aggressive type of skin cancer with increasing incidence rates worldwide. On the other hand, watercress is a rich source of phenethyl isothiocyanate (PEITC), among others, which has been widely investigated for its anticancer properties against various cancers. In the present study, we evaluated the role of a watercress extract in modulating apoptotic induction in an in vitro model of human malignant melanoma consisting of melanoma (A375, COLO-679, COLO-800), non-melanoma epidermoid carcinoma (A431) and immortalized, non-tumorigenic keratinocyte (HaCaT) cells. Moreover, the chemical composition of the watercress extract was characterized through UPLC MS/MS and other analytical methodologies. In addition, cytotoxicity was assessed by the alamar blue assay whereas apoptosis was determined, initially, by a multiplex activity assay kit (measuring levels of activated caspases -3, -8 and -9) as well as by qRT-PCR for the identification of major genes regulating apoptosis. In addition, protein expression levels were evaluated by western immunoblotting. Our data indicate that the extract contains various phytochemicals (e.g. phenolics, flavonoids, pigments, etc.) while isothiocyanates (ITCs; especially PEITC) were the most abundant. In addition, the extract was shown to exert a significant time- and dose-dependent cytotoxicity against all malignant melanoma cell lines while non-melanoma and non-tumorigenic cells exhibited significant resistance. Finally, expression profiling revealed a number of genes (and corresponding proteins) being implicated in regulating apoptotic induction through activation of the intrinsic apoptotic cascade. Overall, our data indicate the potential of PEITC as a promising anti-cancer agent in the clinical management of human malignant melanoma.
摘要：
恶性黑色素瘤是最具侵袭性的皮肤癌类型，在全球范围内发病率不断上升。另一方面,豆瓣菜是异硫氰酸苯乙酯（PEITC）的丰富来源，其中,其针对各种癌症的抗癌特性已被广泛研究。在本研究中,我们评估了豆瓣菜提取物在调节由黑色素瘤组成的人恶性黑色素瘤体外模型中的凋亡诱导中的作用（A375，COLO-679，COLO-800），非黑色素瘤表皮样癌（A431）和永生化，非致瘤性角质形成细胞（HaCaT）细胞。此外，通过UPLCMS/MS和其他分析方法表征豆瓣菜提取物的化学组成。此外,细胞毒性通过Alamar蓝测定法评估，而细胞凋亡测定，最初,通过多重活性测定试剂盒（测量活化的半胱天冬酶-3，-8和-9的水平）以及通过qRT-PCR鉴定调节细胞凋亡的主要基因。此外,蛋白质表达水平通过蛋白质免疫印迹法评估。我们的数据表明，提取物含有各种植物化学物质（例如酚类，黄酮类化合物，颜料,等。)，而异硫氰酸酯(ITC；尤其是PEITC)最丰富。此外,该提取物对所有恶性黑素瘤细胞系均表现出显著的时间和剂量依赖性细胞毒性,而非黑素瘤和非致瘤细胞表现出显著的耐药性.最后,表达谱分析显示，许多基因（和相应的蛋白质）参与通过激活内在凋亡级联反应来调节凋亡诱导。总的来说,我们的数据表明,PEITC在人类恶性黑色素瘤的临床治疗中具有作为一种有前景的抗癌药物的潜力.
