A protocol for isolation and culture of mesenchymal stem cells from mouse compact bone
A protocol for isolation and culture of mesenchymal stem cells from mouse compact bone
复制标题
小鼠致密骨间充质干细胞分离和培养方案
DOI：
10.1038/nprot.2009.238
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发表时间：
2010-01-01
期刊：
影响因子：
14.8
通讯作者：
Mao, Ning
中科院分区：
文献类型：
--
作者：
Zhu, Heng;Guo, Zi-Kuan;Mao, Ning
与人类不同，由于造血细胞污染了培养物，小鼠骨髓来源的间充质干细胞（MSCs）不容易通过粘附在塑料上获得。这里描述的方案设计是基于致密骨骼中大量的间充质干细胞和骨髓腔和骨骼内部界面中存在的造血细胞这一现象。该过程包括将骨髓从长骨中冲洗出来，用II型胶原酶消化骨片，剥夺释放的细胞，培养消化的骨碎片，其中成纤维细胞样细胞在指定的培养基中迁移和生长。整个技术需要5天才能使贴壁细胞顺利传代。进一步鉴定证实这些细胞是间充质干细胞。我们提供了一种简单且可重复的方法来收集小鼠间充质干细胞，不需要通过分选或免疫磁性技术消耗造血细胞。
Unlike humans, mouse bone marrow-derived mesenchymal stem cells (MSCs) cannot be easily harvested by adherence to plastic owing to the contamination of cultures by hematopoietic cells. The design of the protocol described here is based on the phenomenon that compact bones abound in MSCs and hematopoietic cells exist in the marrow cavities and the inner interfaces of the bones. The procedure includes flushing bone marrow out of the long bones, digesting the bone chips with collagenase type II, deprivation of the released cells and culturing the digested bone fragments, out of which fibroblast-like cells migrate and grow in the defined medium. The entire technique requires 5 d before the adherent cells are readily passaged. Further identification assays confirm that these cells are MSCs. We provide an easy and reproducible method to harvest mouse MSCs that does not require depletion of hematopoietic cells by sorting or immunomagnetic techniques.