RANKL诱导小鼠单核细胞RAW264．7分化成成熟破骨细胞

Osteoclastogenesis in RAW264. 7 cells induced by RANKL

目的观察小鼠的单核／巨噬细胞RAW264．7的一般生物学特征及在RANKL诱导下形成成熟破骨细胞的特征。方法 RANKL诱导RAW264．7细胞6d后,用抗酒石酸酸性磷酸酶(TRAP) 染色法观察TRAP阳性多核细胞,吖啶橙染色激光共聚焦显微镜(LCSM)观察多核细胞形态;诱导RAW264．7细胞9d后,RT-PCR检测RAW264．7细胞的破骨细胞表型和功能基因表达及其 RANKL诱导后变化;诱导RAW264．7细胞12d后,钙磷覆盖的破骨细胞活性分析板观察破骨细胞的骨吸收功能。结果 RAW264．7细胞TRAP染色阴性,单核或2个核,能表达破骨细胞表型和功能基因,无骨吸收功能。RANKL可诱导RAW264．7细胞形成TRAP阳性成熟的多核破骨细胞, 上调CathepsinK、CAⅡ、integrinβ3等基因。mRNA的表达。结论 RAW264．7具有破骨细胞特征性基因表达谱,是一种较好的破骨前体细胞模型。RANKL可诱导RAW264．7细胞形成成熟破骨细胞。

Objective To observe the biologic characteristics of RAW264. 7 cell, monocyte/macro-phage cell origined from mouse, and to study the characteristics of mature osteoclast induced by RANKL from RAW264. 7. Methods RAW264. 7 cells were treated with or without RANKL for 6 days, and the morphological change was observed after TRAP staining and by LCSM. Osteoclast phenotypic and functional genes were detected with semiquantitative RT-PCR when RAW264. 7 cells were induced for 9 days. Osteoclastic absorption functions were evaluated by bone resorption pit formation on osteoclast activity analysis platelet when RAW264. 7 cells were induced for 12 days. Results RAW264. 7 cells show TRAP-negative with monocyte or 2 nucleus, expressing osteoclast phenotypic and functional genes without capability of absorbing bone tissue. RAW264. 7 cells could be induced to mature multiple nucleus osteoclasts, The expression of cathepsin K, CAII, integrin β 3 mRNA could be up - regulated by RANKL. Conclusions RAW264. 7 cell could be used as a preferable preoste-oclast model in respect of its osteoclast characteristic gene expression profile. RANKL could induce RAW264. 7 cell to mature osteoclast.