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Cloning and functional analysis of specific molecules of osteoclast

破骨细胞特定分子的克隆及功能分析

基本信息

批准号：
09557086
负责人：
SUDA Toshio
金额：
$ 8.32万
依托单位：
Kumamoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1999
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09557086/
关键词：
Osteoclasts STK osteoblasts TYRO3 c-KitィイD1+ィエD1Mac-1ィイD1dullィエD1c-FmsィイD1+ィエD1 cells RANK M-CSF RANKL KIT Mac-1 c-FMS TRAP ODF RANK STK(RON)ビタミンD3 TYRO3 GAS6

项目摘要

Bone is continuously forming and being resorbed. This process is accomplished by precise coordination of two cell types: osteoblasts, which deposit the calcified bone matrix, and osteoclasts, which derived from hematopoietic stem cells and resorb the bone tissue. Mononuclear osteoclasts differentiate from the same precursor cells as macrophages and fuse with each other to become multinuclear osteoclasts. To clarify the differentiation of osteoclasts from precursors and their relationship to macrophage precursors, we have analyzed the functions of osteoclast-specific genes.At first we have shown that STK and TYRO 3 receptor tyrosine kinases are expressed on osteoclasts and that they are involved in the bone resorption in the presence of each ligand, MSP and GAS6, respectively.Next, by using FACS, we identified c-KitィイD1+ィエD1Mac-1ィイD1dullィエD1c-Fms (M-CSFR)ィイD1+ィエD1 cells in murine bone marrow as precursor cells. C-KitィイD1+ィエD1Mac-1ィイD1dullィエD1c-FmsィイD1+ィエD1 cells differentiate into c- FmsィイD1+ィエD1 cells in 2 days' co-culture with ST-2 stromal cells. We investigated the commitment process of c-KitィイD1+ィエD1Mac-1ィイD1dullィエD1c-FmsィイD1+ィエD1 cells to osteoclasts in the presence of M-CSF and soluble RANK-ligand (sRANKL) instead of stromal cells. M-CSF affect c-KitィイD1+ィエD1Mac-1ィイD1dullィエD1c-FmsィイD1+ィエD1 cells to induce RANK in 24 hrs at mRNA and protein level. These cells can differentiate into osteoclasts in the co-presence of M-CSF and sRANKL. In the presence of only M-CSF, they did differentiate into macrophages. Delayed addition of RANKL revealed that expression of RANK receptor and immediate binding of RANK-ligand is critical for osteoclast differentiation. Thus, the RANK-RANKL system determines the osteoclast differentiation of bipotential precursors in the default pathway of macrophagic differentiation.

骨骼不断形成并被吸收。这一过程是通过两种细胞类型的精确协调来完成的：成骨细胞，沉积钙化骨基质；破骨细胞，源自造血干细胞，吸收骨组织。单核破骨细胞由与巨噬细胞相同的前体细胞分化而来，并相互融合成为多核破骨细胞。为了阐明破骨细胞与前体细胞的分化及其与巨噬细胞前体细胞的关系，我们分析了破骨细胞特异性基因的功能。首先，我们表明 STK 和 TYRO 3 受体酪氨酸激酶在破骨细胞上表达，并且它们分别在每种配体 MSP 和 GAS6 存在的情况下参与骨吸收。接下来，通过使用 FACS，我们鉴定出小鼠骨髓中的c-KitィイD1+ィエD1Mac-1ィイD1dullィエD1c-Fms (M-CSFR)ィイD1+ィエD1细胞作为前体细胞。 C-KitィイD1+ィエD1Mac-1ィイD1dullエD1c-FmsィイD1+ィエD1细胞与ST-2基质细胞共培养2天后分化为c-FmsィイD1+ィエD1细胞。我们研究了在 M-CSF 和可溶性 RANK 配体 (sRANKL) 而不是基质细胞存在的情况下，c-KitィイD1+ィエD1Mac-1ィイD1dullィエD1c-FmsィイD1+ィエD1 细胞向破骨细胞的定型过程。 M-CSF影响c-KitィイD1+ィエD1Mac-1ィイD1dullィエD1c-FmsィイD1+ィエD1细胞在24小时内在mRNA和蛋白质水平上诱导RANK。这些细胞可以在 M-CSF 和 sRANKL 共存的情况下分化为破骨细胞。在仅存在 M-CSF 的情况下，它们确实分化为巨噬细胞。延迟添加 RANKL 揭示了 RANK 受体的表达和 RANK 配体的立即结合对于破骨细胞分化至关重要。因此，RANK-RANKL系统决定了巨噬细胞分化默认途径中双能前体的破骨细胞分化。