Retrovirus-mediated conditional immortalization and analysis of established cell lines of osteoclast precursor cells.
Retrovirus-mediated conditional immortalization and analysis of established cell lines of osteoclast precursor cells.
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DOI：
10.1016/j.bbrc.2006.09.020
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发表时间：
2006-11
影响因子：
3.1
通讯作者：
Shigehisa Kawata;Jun Suzuki;Masahiro Maruoka;Megumi Mizutamari;N. Ishida-Kitagawa;K. Yogo;P. Jat;T. Shishido
中科院分区：
文献类型：
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作者：
Shigehisa Kawata;Jun Suzuki;Masahiro Maruoka;Megumi Mizutamari;N. Ishida-Kitagawa;K. Yogo;P. Jat;T. Shishido
破骨细胞前体细胞（OPCs）以前已经从SV40温度敏感性T抗原表达转基因小鼠的骨髓细胞中建立。在这里，我们使用逆转录病毒介导的基因转移有条件地永生化OPCs表达温度敏感性大T抗原（tsLT）从野生型骨髓细胞。永生化的OPCs在33.5°C的允许温度下增殖，但在39°C的非允许温度下停止生长。在NFκB受体激活因子配体（RANKL）存在下，OPCs在33.5°C下分化为抗酒石酸酸性磷酸酶（TRAP）阳性细胞，并形成多核破骨细胞。从这些OPC中，我们克隆了两种类型的细胞系。两者均分化为TRAP阳性细胞，但一个形成多核破骨细胞，而另一个在RANKL存在下保持未融合。这些结果表明，建立的细胞系是有用的分化机制的分析，特别是多核破骨细胞的形成。逆转录病毒介导的条件永生化是一种从原代骨髓细胞获得OPCs的有效方法。
Osteoclast precursor cells (OPCs) have previously been established from bone marrow cells of SV40 temperature-sensitive T antigen-expressing transgenic mice. Here, we use retrovirus-mediated gene transfer to conditionally immortalize OPCs by expressing temperature-sensitive large T antigen (tsLT) from wild type bone marrow cells. The immortalized OPCs proliferated at the permissive temperature of 33.5°C, but stopped growing at the non-permissive temperature of 39°C. In the presence of receptor activator of NFκB ligand (RANKL), the OPCs differentiated into tartrate-resistant acid phosphatase (TRAP)-positive cells and formed multinucleate osteoclasts at 33.5°C. From these OPCs, we cloned two types of cell lines. Both differentiated into TRAP-positive cells, but one formed multinucleate osteoclasts while the other remained unfused in the presence of RANKL. These results indicate that the established cell lines are useful for analyzing mechanisms of differentiation, particularly multinucleate osteoclast formation. Retrovirus-mediated conditional immortalization should be a useful method to immortalize OPCs from primary bone marrow cells.