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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情况。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。
BACKGROUND: Osteoclasts, as a terminal cell, are difficult to obtain, and lack of mature osteoclast lines and other factors have limited their application. Previous domestic and foreign acquisition of osteoclasts generally use stromal cell induction culture or co-culture, or the use of nuclear factor kappa B receptor activating factor ligand and macrophage colony stimulating factor together to induce the formation of mature osteoclasts. OBJECTIVE: To observe the general biological characteristics of mouse RAW264.7 monocyte-macrophage cells and to analyze the feasibility of forming mature osteoclasts under the induction of NF-κB ligand. Methods: RAW264.7 cells were induced with RAW264.7 cells after RAW264.7 cells were cultured for 7 days. The results of tartrate-resistant acid phosphatase staining were positive, tartrate-resistant acid phosphatase staining was positive, and the number of nuclei ≥3 cell. The fibrin actin ring was observed by phalloidin fluorescent staining, and the absorption lacunae on the surface of bovine bone were observed by toluidine blue staining. RESULTS AND CONCLUSION: NF-κB receptor activator ligand induces RAW264.7 cells to form antinuclear acid phosphatase-positive multinucleate cells, forming a fibrinous actin loop. Electron microscopy shows the discs are round or oval Absorb dimples. Tip RAW264.7 is a better model of osteoclast precursor cells can be used for osteoclast differentiation studies. Inducing the differentiation and maturation of RAW264.7 cells by the single nuclear factor kappa B receptor activating factor ligand reduces the application of macrophage colony stimulating factor and makes the culture system simple and easy to operate. The induced cells have high purity and are suitable for osteoclast Cell biology and biochemistry research.