Department of Nutritional Science and Food Management, Ewha Womans University, 11-1 Daehyun-dong, Seodaemun-gu, Seoul 120-750, Korea.
Nutr Res Pract. 2010 Aug;4(4):276-82. doi: 10.4162/nrp.2010.4.4.276. Epub 2010 Aug 31.
The principal objective of this study was to evaluate the chemopreventive and therapeutic effects of a combination of all-trans-retinoic acid (RA) and knockdown of delta-like 1 homologue (Drosophila) (DLK1) on neuroblastoma, the most common malignant disease in children. As unfavorable neuroblastoma is poorly differentiated, neuroblastoma cell was induced differentiation by RA or DLK1 knockdown. Neuroblastoma cells showed elongated neurite growth, a hallmark of neuronal differentiation at various doses of RA, as well as by DLK1 knockdown. In order to determine whether or not a combination of RA and DLK1 knockdown exerts a greater chemotherapeutic effect on neuroblastoma, cells were incubated at 10 nM RA after being transfected with SiRNA-DLK1. Neuronal differentiation was increased more by a combination of RA and DLK1 knockdown than by single treatment. Additionally, in order to assess the signal pathway of neuroblastoma differentiation induced by RA and DLK1 knockdown, treatment with the specific MEK/ERK inhibitors, U0126 and PD 98059, was applied to differentiated neuroblastoma cells. Differentiation induced by RA and DLK1 knockdown increased ERK phosphorylation. The MEK/ERK inhibitor U0126 completely inhibited neuronal differentiation induced by both RA and DLK1 knockdown, whereas PD98059 partially blocked neuronal differentiation. After the withdrawal of inhibitors, cellular differentiation was fully recovered. This study is, to the best of our knowledge, the first to demonstrate that the specific inhibitors of the MEK/ERK pathway, U0126 and PD98059, exert differential effects on the ERK phosphorylation induced by RA or DLK1 knockdown. Based on the observations of this study, it can be concluded that a combination of RA and DLK1 knockdown increases neuronal differentiation for the control of the malignant growth of human neuroblastomas, and also that both MEK1 and MEK2 are required for the differentiation induced by RA and DLK1 knockdown.
本研究的主要目的是评估全反式维甲酸（RA）与下调 delta 样 1 同源物（果蝇）（DLK1）联合应用对神经母细胞瘤（儿童最常见的恶性疾病）的化学预防和治疗作用。由于不良的神经母细胞瘤分化不良，通过 RA 或 DLK1 敲低诱导神经母细胞瘤细胞分化。RA 以及 DLK1 敲低在不同剂量下，神经母细胞瘤细胞表现出长突起生长，这是神经元分化的标志。为了确定 RA 和 DLK1 敲低的联合应用是否对神经母细胞瘤具有更大的化疗作用，在用 SiRNA-DLK1 转染后，将细胞在 10 nM RA 中孵育。RA 和 DLK1 敲低的联合作用比单一处理更能增加神经元分化。此外，为了评估 RA 和 DLK1 敲低诱导的神经母细胞瘤分化的信号通路，用特异性 MEK/ERK 抑制剂 U0126 和 PD98059 处理分化的神经母细胞瘤细胞。RA 和 DLK1 敲低诱导的分化增加了 ERK 磷酸化。MEK/ERK 抑制剂 U0126 完全抑制了 RA 和 DLK1 敲低诱导的神经元分化，而 PD98059 部分阻断了神经元分化。在抑制剂撤出后，细胞分化完全恢复。据我们所知，这项研究首次表明，MEK/ERK 通路的特异性抑制剂 U0126 和 PD98059 对 RA 或 DLK1 敲低诱导的 ERK 磷酸化产生不同的影响。基于本研究的观察结果，可以得出结论，RA 和 DLK1 敲低的组合增加了神经元分化，以控制人类神经母细胞瘤的恶性生长，并且 MEK1 和 MEK2 都需要 RA 和 DLK1 敲低诱导的分化。
