Retinoic acid-induced neuritogenesis of human neuroblastoma SH-SY5Y cells is ERK independent and PKC dependent
Retinoic acid-induced neuritogenesis of human neuroblastoma SH-SY5Y cells is ERK independent and PKC dependent
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DOI：
10.1002/jnr.10848
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发表时间：
2004-01-15
影响因子：
4.2
通讯作者：
Tredici, G
中科院分区：
文献类型：
--
作者：
Miloso, M;Villa, D;Tredici, G
维甲酸（RA）是维生素A的活性代谢产物，是一种参与神经系统发育和分化的天然形态原。为了阐明RA诱导的轴突发生中涉及的信号机制，我们使用人类神经母细胞瘤SH-SY 5 Y细胞（一种用于研究RA作用的体外模型）来研究细胞外信号调节激酶（ERK）1和2在RA诱导的轴突发生中的作用。和细胞生存。从免疫印迹实验中，我们观察到，RA诱导延迟，但持续的ERK 1和ERK 2磷酸化（直到96小时），显着减少特定的丝裂原活化蛋白激酶（MAPK）/ERK激酶（MEK）抑制剂U 0126。对于随后的研究，我们选择24小时作为参考时间。ERK激活的抑制并不影响RA诱导的神经突发生（神经突承载细胞和神经突长度的百分比），但显着降低细胞存活。此外，我们分析了介导ERK激活的信号通路。我们的研究结果表明，RA诱导的ERK磷酸化不遵循经典的Raf激酶依赖性途径。蛋白激酶C（PKC）和磷脂酰肌醇3-激酶（PI 3-K）是参与ERK信号通路的可能的替代激酶。事实上，在存在特异性PKC抑制剂GF 109203 X或特异性PI 3-K抑制剂渥曼青霉素的情况下，我们观察到ERK磷酸化的显著剂量依赖性降低。GF 109203 X以浓度依赖性方式降低RA诱导的轴突发生和细胞存活。这些结果表明，在RA诱导的轴突发生的早期阶段发挥重要作用的是PKC，而不是ERK 1和ERK 2。(C)2003 Wiley-Liss，Inc.
Retinoic acid (RA), an active metabolite of vitamin A, is a natural morphogen involved in development and differentiation of the nervous system. To elucidate signaling mechanisms involved in RA-induced neuritogenesis, we used human neuroblastoma SH-SY5Y cells, an established in vitro model for studying RA action, to examine the role of extracellular signal-regulated kinase (ERK) 1 and 2 in RA-induced neuritogenesis and cell survival. From immunoblotting experiments, we observed that RA induced delayed but persistent ERK1 and ERK2 phosphorylation (until 96 hr) that was reduced significantly by the specific mitogen-activated protein kinase (MAPK)/ERK kinase (MEK) inhibitor U0126. For the subsequent studies we chose 24 hr as the reference time. Inhibition of ERK activation did not affect RA-induced neuritogenesis (percentage of neurite-bearing cells and neurite length) but significantly reduced cell survival. In addition we analyzed the signaling pathway that mediates ERK activation. Our results suggest that RA-induced ERK phosphorylation does not follow the classic Raf kinase-dependent pathway. Protein kinase C (PKC) and phosphatidylinositol 3-kinase (PI 3-K) are possible alternative kinases involved in the ERK signaling pathway. In fact, in the presence of the specific PKC inhibitor GF 109203X, or the specific PI 3-K inhibitor wortmannin, we observed a significant dose-dependent reduction in ERK phosphorylation. RA-induced neuritogenesis and cell survival were reduced by GF 109203X in a concentration-dependent manner. These results suggest that rather than ERK1 and ERK2, it is PKC that plays an important role during early phases of RA-induced neuritogenesis. (C) 2003 Wiley-Liss, Inc.