NMDA glutamate receptors are expressed by osteoclast precursors and involved in the regulation of osteoclastogenesis
NMDA glutamate receptors are expressed by osteoclast precursors and involved in the regulation of osteoclastogenesis
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DOI：
10.1002/jcb.10625
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发表时间：
2003-10-01
影响因子：
4
通讯作者：
Chenu, C
中科院分区：
文献类型：
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作者：
Merle, B;Itzstein, C;Chenu, C
我们之前在成熟破骨细胞中鉴定出功能性 N-甲基-D-天冬氨酸 (NMDA) 谷氨酸受体，并证明它们参与体外骨吸收。在目前的工作中，我们使用两种体外模型（小鼠骨髓单核细胞RAW 264.7细胞系和小鼠骨髓细胞）研究了破骨细胞前体的NMDA受体（NMDAR）表达及其在破骨细胞生成中的作用，这两种模型在巨噬细胞集落刺激因子（M-CSF）和Rank配体（RankL）存在下分化为破骨细胞。使用特定探针的 RT-PCR 分析，我们发现 RAW 264.7 细胞和小鼠骨髓细胞表达 NMDAR 亚基 NMDA 受体 1 (NR1) 和 NMIDA 受体 2 (NR2) A、B 和 D 的 mRNA。这些亚基在从未分化前体到成熟再吸收破骨细胞的分化序列中一直表达。半定量PCR分析显示在分化过程中这些亚基的表达没有受到调节。 NMDAR 的两种特异性非竞争性拮抗剂 MK801 和 DEP 在两种模型中均以剂量依赖性方式抑制破骨细胞形成，表明破骨细胞生成需要激活破骨细胞前体表达的 NMDAR。仅在培养的前 2 天添加 MK801 没有效果，表明 NMDAR 参与破骨细胞形成的后期阶段。最后，我们使用蛋白质印迹和免疫荧光证明，特定激动剂在 RAW 264.7 细胞中激活 NMDAR 会诱导 NF-κ 13 的核转位，这是破骨细胞形成所需的因子。总而言之，我们的结果表明破骨细胞前体表达 NMDAR，通过激活 NF-κ B 途径参与破骨细胞分化过程。 (C) 2003 Wiley-Liss, Inc.
We previously identified functional N-methyl-D-aspartate (NMDA) glutamate receptors in mature osteoclasts and demonstrated that they are involved in bone resorption in vitro. In the present work, we studied the expression of NMDA receptors (NMDAR) by osteoclast precursors and their role in osteoclastogenesis using two in vitro models, the murine myelomonocytic RAW 264.7 cell line and mouse bone marrow cells, both of which differentiate into osteoclasts in the presence of macrophage colony-stimulating factor (M-CSF) and Rank ligand (RankL). Using RT-PCR analysis with specific probes, we showed that RAW 264.7 cells and mouse bone marrow cells express mRNA of NMDAR subunits NMDA receptor 1 (NR1) and NMIDA receptor 2 (NR2) A, B, and D. These subunits are expressed all along the differentiation sequence from undifferentiated precursors to mature resorbing osteoclasts. Semi-quantitative PCR analysis showed no regulation of the expression of these subunits during the differentiation process. Two specific non competitive antagonists of NMDAR, MK801 and DEP, dose-dependently inhibited osteoclast formation in both models, indicating that osteoclastogenesis requires the activation of NMDAR expressed by osteoclast precursors. MK801 had no effect when added only during the first 2 days of culture, suggesting that NMDAR are rather involved in the late stages of osteoclast formation. Finally, we demonstrated using Western-blotting and immunofluorescence that activation of NMDAR in RAW 264.7 cells by specific agonists induces nuclear translocation of NF-kappa 13, a factor required for osteoclast formation. Altogether, our results indicate that osteoclast precursors express NMDAR that are involved in the osteoclast differentiation process through activation of the NF-kappa B pathway. (C) 2003 Wiley-Liss, Inc.