中国现代医学杂志2017，Vol.27Issue(18)：1-6,6.DOI:10.3969/j.issn.1005-8982.2017.18.001
大鼠骨髓单核细胞的快速分离方法及向破骨细胞的诱导分化
A method about isolation and cultivation of rat bone marrow monocytes and differentiation into osteoclasts
摘要
Abstract
Objective To establish a simple and efficient method to isolate the rat bone marrow monocytes in vitro,observe the characteristics of growth and induce the differentiation of osteoclasts.Methods The tibia and femur of SD rats were isolated under aseptic condition.The bone marrow tissues were separated by EP tube and the transfer gun head.Red blood cell lysate was used to remove the red blood cells.The suspended cells were collected after cell culture overnight.Then M-CSF was added to culture the cells and adherent bone marrow monocytes were obtained.The growth process of bone marrow monocytes was morphologically observed.The cell growth curve was measured by CCK-8.Flow cytometry was used to detect CD11b expression on cell surface.After differentiation of the monocytes by adding M-CSF and RANKL,TRAP staining and CTR immunofluorescence staining were used to identify whether they can differentiate to mature osteoclasts.Results The rat bone marrow monocytes obtained after culturing for 1 day by these methods showed small circular shape with few mixed cells.After 3 days,the number of the cells increased slightly and the antennae began to appear at both sides.After 5 days,the cells increased significantly and were in oval shape,the antennae at both sides were obvious.The cell proliferation depended on M-CSF.Flow cytometry showed that monocytes obtained had high purity by this method.TRAP staining and CTR immunofluorescence staining results showed that monocytes obtained by this method could be successfully induced to mature osteoclasts.Conclusions Monocytes can be isolated and obtained rapidly by this method.The phenotype of the obtained cells is stable and suitable for further study on bone metabolic diseases.关键词
分离培养方法/破骨细胞/骨髓单核细胞/分化/骨代谢疾病Key words
isolation and culture method/osteoclast/bone marrow monocyte/differentiation/bone metabolic disease分类
生物科学引用本文复制引用
王礼宁,马勇,郑苏阳,郭杨,袁翰,孙杰..大鼠骨髓单核细胞的快速分离方法及向破骨细胞的诱导分化[J].中国现代医学杂志,2017,27(18):1-6,6.基金项目
国家自然科学基金面上项目(No:81473692) （No:81473692）
江苏省自然科学基金青年基金项目(No:BK20151007) （No:BK20151007）
