Sato Kazuomi, Ando Ryosuke, Kobayashi Honoka, Nishio Takashi
Graduate School of Agriculture, Tamagawa University, Machida, Tokyo, 194-8610, Japan.
Department of Life Science, College of Agriculture, Tamagawa University, Machida, Tokyo, 194-8610, Japan.
Mol Cell Biochem. 2016 Dec;423(1-2):39-52. doi: 10.1007/s11010-016-2823-x. Epub 2016 Sep 16.
Non-steroidal anti-inflammatory drugs are frequently used for the treatment of inflammation, pain, and fever. In this study, we found that 2-ethoxybenzamide (ETZ) significantly enhanced melanin synthesis in B16F1 melanoma cells, and also induced melanosome formation. Therefore, we investigated the mechanism by which ETZ up-regulated melanin synthesis. Western blot analysis demonstrated that ETZ increased melanogenic protein levels, except that for TRP-2. Moreover, semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and real-time RT-PCR analyses showed that ETZ enhanced the mRNA levels of melanogenic genes, including microphthalmia-associated transcription factor and melanocortin 1 receptor. We also observed phosphorylation of cAMP response element-binding protein (CREB) following ETZ treatment. However, ETZ did not affect intracellular cAMP levels. ERK was also activated by ETZ treatment, and melanin content was enhanced upon treatment with the specific ERK inhibitor PD98059. Together, our results indicate that ETZ induces melanin synthesis via CREB phosphorylation.
非甾体抗炎药常用于治疗炎症、疼痛和发热。在本研究中，我们发现2-乙氧基苯甲酰胺（ETZ）显著增强了B16F1黑色素瘤细胞中的黑色素合成，并且还诱导了黑素小体的形成。因此，我们研究了ETZ上调黑色素合成的机制。蛋白质免疫印迹分析表明，ETZ增加了黑色素生成蛋白的水平，但酪氨酸酶相关蛋白-2（TRP-2）除外。此外，半定量逆转录-聚合酶链反应（RT-PCR）和实时RT-PCR分析表明，ETZ增强了包括小眼相关转录因子和黑皮质素1受体在内的黑色素生成基因的mRNA水平。我们还观察到ETZ处理后cAMP反应元件结合蛋白（CREB）的磷酸化。然而，ETZ不影响细胞内cAMP水平。ETZ处理也激活了细胞外信号调节激酶（ERK），并且用特异性ERK抑制剂PD98059处理后黑色素含量增加。总之，我们的结果表明ETZ通过CREB磷酸化诱导黑色素合成。
