核因子κB受体活化因子配体诱导形成的成熟破骨细胞

背景：破骨细胞作为一种终末细胞，获取困难，而且没有成熟破骨细胞株等因素限制了其应用。先前国内外对破骨细胞的获取一般采用基质细胞诱导培养或共培养，或运用核因子κB受体活化因子配体和巨噬细胞集落刺激因子共同作用诱导形成成熟的破骨细胞。目的：观察小鼠单核巨噬细胞RAW264．7的一般生物学特征，分析其在核因子κB受体活化因子配体诱导下形成成熟破骨细胞的可行性。方法：培养RAW264．7后，用核因子κB受体活化因子配体诱导RAW264．7细胞7d后观察抗酒石酸酸性磷酸酶染色结果，以抗酒石酸酸性磷酸酶染色阳性，细胞核≥3个为破骨细胞。以鬼笔环肽荧光染色观察纤维性肌动蛋白环，甲苯胺蓝染色观察牛骨片表面的吸收陷窝隋况。结果与结论：核因子κB受体活化因子配体可诱导RAW264．7细胞形成抗酒石酸酸性磷酸酶染色阳性的多核细胞，形成纤维性肌动蛋白环，电镜下可见骨片上圆形或椭圆形的吸收陷窝。提示RAW264．7是一种较好的破骨前体细胞模型，可用于破骨细胞分化研究。单用核因子κB受体活化因子配体诱导RAW264．7细胞分化成熟，减少了巨噬细胞集落刺激因子的应用，使培养体系更加简单，易于操作，诱导出的细胞纯度高，适合于破骨细胞的生物学和生化研究。

Receptor activator of nuclear factor kappa B ligand-induced mature osteoclasts

BACKGROUND： Osteoclast as a terminal cell is difficult to obtain, and the shortage of mature osteoclaststrains limits its application. Previous studies have shown that stromal cell culture or co-culture is commonly used to obtain the osteoclasts, and the receptor activator of nuclear factor κB ligand and macrophage colony-stimulating factor can be used in combination in order to obtain the mature osteoclasts. OBJECTIVE： To observe the biological characteristics of mouse monocyte macrophage RAW264.7, and to analyze the feasibility of receptor activator of nuclear factor κB ligand-mediated differentiation of monocyte macrophage RAW264.7 into osteoclasts. METHODS： The RAW264.7 cells were cultured, and then the RAW264.7 cells were induced with receptor activator of nuclear factor κB ligand for 7 days to observe the tartrate-resistant acid phosphatase staining results. The positive tartrate-resistant acid phosphatase staining and nucleus 〉 3 were considered as the osteoclasts. Phalloidin staining was used to observe fibrous actin ring, and toluidine blue staining was used to observe the resorption pits on the surface of bovine bone slice. RESULTS AND CONCLUSION： RAW264.7 cells could be induced into tartrate-resistant acid phosphatase-positive multinuclear cells with receptor activator of nuclear factor κB ligand, and formed fibrous actin rings; round or ellipse bone resorption pits were found in bone slice surface by microscope. RAW264.7 cells were considered as a good preosteoclast model that used for research of osteoclast differentiation. Receptor activator of nuclear factor κB ligand caninduce the differentiation and maturation of RAW264.7 cells, reduce the application of macrophage colony-stimulating factor and make the training system simpler and easier to operate, and the induced cells have the high purity suitable for biological and biochemical studies of osteoclasts.