胰酶消化法纯化诱导不同年龄段SD大鼠骨髓破骨细胞的体外培养
Isolation of Bone Marrow-derived Rat Osteoclast-like Cells with the Digestion of Trypsin
作者：董强;梁星;陈悦;徐凌;张庆鸿;夏露;陈明;付俊
Author：DONG Qiang1, LIANG Xing2, CHEN Yue2, XU Ling1, ZHANG Qing-hong1, XIA Lu1, CHEN Ming1, FU Jun1
收稿日期：2006-08-25 年卷（期）页码：2006,24(04):350-
期刊名称：华西口腔医学杂志
Journal Name：West China Journal of Stomatology
关键字：破骨细胞,胰酶消化,培养,纯化,
Key words：osteoclast,trypsin digestion,culture,purify,
基金项目：
国家自然科学基金资助项目（30271428）；教育部“优秀青年教师资助计划”资助项目[教人司（2002）40号]；高等学校博士学科点专项科研基金资助项目（B1200058）
中文摘要
目的 用胰酶消化法纯化获取大量高纯度破骨细胞，为进行相关分子生物学研究奠定基础。方法 以1,25-（OH）2D3 /地塞米松诱导培养不同年龄段（1、12、24、36 d）SD大鼠的骨髓破骨细胞，采用0.25%胰蛋白酶/0.02%乙二胺四乙酸纯化。通过倒置相差显微镜观察细胞形态，采用抗酒石酸酸性磷酸酶（Trap）染色和扫描电镜观察鉴定破骨细胞，并计数分析。结果 1、12、24、36 d组大鼠骨髓破骨细胞均培养成功，经Trap染色和扫描电镜观察证实为体外有噬骨能力的破骨细胞，纯化率达到90％。1 d和12 d组破骨细胞出现较早且数量较多，细胞计数两组间无统计学差异（P>0.05），但它们与其余各组均有统计学差异（P
英文摘要
Objective To obtain highly enriched osteoclasts in vitro. Methods The bone marrow cells of 1-day, 12-day, 24-day and 36-day old Sprague-Dawley（SD） rats were separately cultured with the presence of 1,25-（OH）2D3 and dexamethasone, and the osteoclast-like cells were identified by Trap staining and scanning electron microscope observation and were purified by trypsin digestion. The count of Trap positive osteoclast-like cells was analyzed statistically. Results The osteoclast-like cells of each group were Trap positive cells and could form the bone absorption lacunas in vitro. The amounts of osteoclast-like cells were different statistically between the groups（P0.05）. Highly enriched teoclast-like cells were harvested by the digestion of 0.25% trypsin and 0.02% EDTA. Conclusion It′s indicated that a large amount of highly enriched osteoclast-like cells could be obtained through the culture of bone marrow cells of 10-day and 12-day old SD rats with 1,25-（OH）2D3 and dexamethasone and the digestion of trypsin/EDTA.
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