RAW 264.7细胞与骨髓源巨噬细胞向破骨细胞分化特性的比较

Comparison between bone marrow-derived macrophages and RAW 264.7 cells in inducing osteoclast differentiation

目的比较RAW 264.7细胞与骨髓源巨噬细胞(bone marrow-derived macrophages,BMMs)向破骨细胞分化的特性。方法骨髓原始细胞经巨噬细胞集落刺激因子(macrophage colony-stimulating factor,M-CSF)刺激3 d,免疫荧光鉴定是否为BMMs;RAW 264.7细胞与BMMs经核因子κB受体活化因子配体(receptor activator of nuclear factorκB ligand,RANKL)诱导4 d后,通过抗酒石酸酸性磷酸酶(tartrate-resistant acid phosphatase,TRAP)染色和纤维肌动蛋白环染色鉴定破骨细胞,并比较两者诱导破骨细胞的效率,同时通过骨板吸收实验检测破骨细胞的骨吸收活性。结果骨髓原始细胞经M-CSF刺激3 d后可得BMMs;RAW 264.7细胞与BMMs经RANKL诱导4 d后均可出现大量TRAP阳性多核破骨细胞且形成纤维肌动蛋白环,二者诱导破骨细胞的效率差异无统计学意义;骨板吸收实验显示RAW 264.7细胞与BMMs骨板上均形成较多的骨吸收瘢痕,且二者的骨吸收活性差异无统计学意义。结论 RAW 264.7细胞与BMMs向破骨细胞分化的特性无明显差异。

Objective The objective of this study was to compare the characteristics between bone marrow-derived macrophage and RAW264.7cells in inducing osteoclast differentiation.Methods Primary bone marrow cells were stimulated by macrophage colony-stimulating factor(M-CSF)for3days,and CD11b immunostaining was used to identify BMMs.RAW264.7cells and BMMs were induced by receptor activator of nuclear factorκB ligand(RANKL)for4days,then tartrate-resistant acid phosphatase(TRAP)staining and F-actin ring staining were used to identify osteoclasts.The efficiency of these two methods for inducing osteoclast were compared.The Osteo Assay Surface Plate was used to measure the bone absorption activities of osteoclasts.Results Primary bone marrow cells were stimulated into BMMs after being exposed to M-CSF for3days.Both RAW264.7cells and BMMs differentiated into TRAP-positive osteoclast following RANKL treatment and formed F-actin rings,and there was no statistical difference between two groups in the efficiency of inducing osteoclast.The Osteo Assay Surface Plate experiment showed that both RAW264.7cells and BMMs formed much more bone absorption scar on the surface of the Osteo Assay Surface Plate,and there was no statistical difference in bone absorption activity between these two.Conclusions There is no significant difference between RAW264.7cells and BMMs in the differentiation of osteoclasts.