Ma Yi-Shih, Hsiao Yung-Ting, Lin Jen-Jyh, Liao Ching-Lung, Lin Chin-Chung, Chung Jing-Gung
School of Chinese Medicine for Post-Baccalaureate, I-Shou University, Kaohsiung, Taiwan, R.O.C.
Department of Chinese Medicine, E-Da Hospital, Kaohsiung, Taiwan, R.O.C.
Anticancer Res. 2017 Nov;37(11):6223-6234. doi: 10.21873/anticanres.12073.
BACKGROUND/AIM: Numerous evidence has shown that PEITC and BITC inhibit cancer cell migration and invasion. In this study, we investigated the anti-metastatic mechanisms of PEITC and BITC in human melanoma cancer A375.S2 cells in vitro.
We used a cell viability assay, an in-vitro scratch wound healing assay, a transwell assay for cell migration and invasion, a gelatin zymography assay, western blotting and EMSA to examine the anti-metastatic mechanisms of PEITC and BITC in A375.S2 cells.
Sublethal concentrations of PEITC (0, 1, 2 and 2.5 μM) and BITC (0, 0.5, 1 and 2 μM) inhibited mobility, migration and invasion of A375.S2 cells that were assayed by wound healing and Transwell filter. PEITC and BITC inhibited MMP-2 activity in A375.S2 cells, as assessed by gelatin zymography assay. Results from western blotting indicated that PEITC (2.5 μM) and BITC (2 μM) decreased the levels of p-p38 following 24 and 48 h treatment. PEITC (1-2.5 μM) reduced the levels of p-JNK1/2 proteins following 48-h treatment but BITC increased p-JNK1/2 levels following 24-h treatment. PEITC (2.5 μM) reduced the levels of p-ERK1/2 proteins following 48-h treatment but BITC (0.5-2 μM) increased p-ERK1/2 levels following 24- and 48-h treatment. PEITC and BITC affect cell migration and invasion of A375.S2 cells via MAPK pathway. PEITC and BITC inhibited MMP-2 activity. PEITC increased NF-κB expression but BITC decreased NF-κB expression in the nucleus. Furthermore, NF-κB p65 binding to DNA was decreased following 2.5 μM PEITC treatment, but increased following treatment with 1-2 μM. However, 0.5-2 μM BITC treatment decreased the binding of NF-κB to DNA in A375.S2 cells, as assessed by electrophoretic mobility shift (EMSA) assay.
Based on these observations, we suggest that PEITC and BITC can be used as anti-metastastic agents of human melanoma cells in the future.
背景/目的：大量证据表明，异硫氰酸苯乙酯（PEITC）和异硫氰酸苄酯（BITC）可抑制癌细胞的迁移和侵袭。在本研究中，我们在体外研究了PEITC和BITC对人黑色素瘤A375.S2细胞的抗转移机制。
我们使用细胞活力测定、体外划痕伤口愈合测定、用于细胞迁移和侵袭的Transwell测定、明胶酶谱测定、蛋白质印迹法和电泳迁移率变动分析（EMSA）来研究PEITC和BITC在A375.S2细胞中的抗转移机制。
亚致死浓度的PEITC（0、1、2和2.5 μM）和BITC（0、0.5、1和2 μM）通过伤口愈合和Transwell滤膜测定法抑制了A375.S2细胞的运动性、迁移和侵袭。通过明胶酶谱测定评估，PEITC和BITC抑制了A375.S2细胞中基质金属蛋白酶-2（MMP-2）的活性。蛋白质印迹法结果表明，在处理24小时和48小时后，PEITC（2.5 μM）和BITC（2 μM）降低了磷酸化p38的水平。在处理48小时后，PEITC（1 - 2.5 μM）降低了磷酸化JNK1/2蛋白的水平，但在处理24小时后，BITC增加了磷酸化JNK1/2的水平。在处理48小时后，PEITC（2.5 μM）降低了磷酸化ERK1/2蛋白的水平，但在处理24小时和48小时后，BITC（0.5 - 2 μM）增加了磷酸化ERK1/2的水平。PEITC和BITC通过丝裂原活化蛋白激酶（MAPK）途径影响A375.S2细胞的迁移和侵袭。PEITC和BITC抑制MMP-2活性。PEITC增加了细胞核中核因子κB（NF-κB）的表达，但BITC降低了其表达。此外，在2.5 μM PEITC处理后，NF-κB p65与DNA的结合减少，但在1 - 2 μM处理后增加。然而，通过电泳迁移率变动分析（EMSA）测定评估，0.5 - 2 μM BITC处理降低了A375.S2细胞中NF-κB与DNA的结合。
基于这些观察结果，我们建议未来PEITC和BITC可作为人黑色素瘤细胞的抗转移剂。
