U0126 and PD98059, specific inhibitors of MEK, accelerate differentiation of RAW264.7 cells into osteoclast-like cells
U0126 and PD98059, specific inhibitors of MEK, accelerate differentiation of RAW264.7 cells into osteoclast-like cells
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DOI：
10.1074/jbc.m208284200
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发表时间：
2002-12-06
影响因子：
4.8
通讯作者：
Nakayama, K
中科院分区：
文献类型：
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作者：
Hotokezaka, H;Sakai, E;Nakayama, K
破骨细胞是从造血细胞分化而来的多核细胞，具有骨吸收的特征。为了研究丝裂原活化蛋白激酶（MAPK）在小鼠单核细胞系RAW 264.7的破骨细胞生成中的作用，我们用p38 MAPK的特异性抑制剂PD 169316和SB 203580处理细胞，以及MAPK细胞外信号调节激酶（ERK）激酶（MEK）的特异性抑制剂，U 0126和PD 98059。当细胞以标准细胞密度（2000-4000个细胞/孔（96孔））铺板时，每种抑制剂阻断向破骨细胞样细胞的分化。然而，MEK抑制剂对破骨细胞生成的影响根据培养过程中的初始细胞密度而变化，因为细胞生长明显受到它们的抑制。当细胞以每孔超过8000个细胞铺板时，观察到分化的显著增强和加速。此外，免疫印迹分析显示，ERK的磷酸化增加了与p38抑制剂的治疗，而MEK抑制剂增加p38的磷酸化，这意味着一个跷跷板样的平衡ERK和p38磷酸化。我们认为，破骨细胞的调节下ERK和p38途径之间的平衡，MEK/ERK途径负调控破骨细胞的生成，而p38途径这样做积极。这是第一个报告，信号转导抑制剂增强破骨细胞。
Osteoclasts are multinucleated cells that differentiate from hematopoietic cells and possess characteristics responsible for bone resorption. To study the involvement of mitogen-activated protein kinases (MAPKs) in osteoclastogenesis of the murine monocytic cell line RAW264.7, which can differentiate into osteoclast-like cells in the presence of the receptor activator of nuclear factor kappa B ligand (RANKL), we treated the cells with specific inhibitors of p38 MAPK, PD169316 and SB203580, and specific inhibitors of MAPK extracellular signaling-regulated kinase (ERK) kinase (MEK), U0126 and PD98059. Each inhibitor blocked differentiation into osteoclast-like cells when the cells were plated at the standard cell density (2000-4000 cells per well (96-well)). However, the effect of MEK inhibitors on osteoclastogenesis varied according to the initial cell density during culture, because cell growth was clearly inhibited by them. When the cells were plated at more than 8000 cells per well, marked enhancement and acceleration of the differentiation were observed. In addition, immunoblot analysis revealed that phosphorylation of ERK was increased by treatment with the p38 inhibitors, whereas the MEK inhibitors increased phosphorylation of p38, which implies a seesaw-like balance between ERK and p38 phosphorylation. We suggest that osteoclastogenesis is regulated under a balance between ERK and p38 pathways and that the MEK/ERK pathway negatively regulates osteoclastogenesis while the p38 pathway does so positively. This is the first report that an inhibitor of signal transduction enhanced osteoclastogenesis.