《中国康复理论与实践》

《中国康复理论与实践》 ›› 2006, Vol. 12 ›› Issue (08): 682-684.

• 基础研究 • 上一篇    下一篇

小鼠骨髓间充质干细胞的分离及其诱导定向脂肪细胞分化

焦嫦亮1,2; 张阳德2; 谢祁阳1,3; 潘一峰2   

  1. 1.海南省干细胞研究所,海南海口市 570102;2.中南大学生物医学工程研究院,湖南长沙市 410078;3.中南大学湘雅医学院生理学系,湖南长沙市 410078
  • 收稿日期:2006-04-24 出版日期:2006-08-01 发布日期:2006-08-01

Separateness and Directional Adipocyte Differentiation of Murine Bone Marrow Mesenchymal Stem Cells

JIAO Chang-liang,ZHANG Yang-de, XIE Qi-yang et al   

  1. The Stem Cells Institute of Hainan Province, Haikou 570102, Hainan, China
  • Received:2006-04-24 Published:2006-08-01 Online:2006-08-01

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被引次数

8

摘要： 目的建立小鼠骨髓间充质干细胞(MSCs)的分离和鉴定方法,稳定和优化骨髓MSCs在体外定向向脂肪细胞分化的适宜诱导条件。方法采取贴壁细胞分离法分离和纯化小鼠骨髓MSCs;选用第5代MSCs分别采用CFUf培养法和CFUf集落再克隆生成CFUf集落法鉴定MSCs的增殖和自我更新能力;利用不同组合的成脂培养基定向诱导MSCs分化为脂肪细胞;利用油红O免疫组化染色鉴定分化后的细胞。结果分离和纯化的小鼠骨髓MSCs呈均匀有序的成纤维母细胞样形态;植入2×103～1×105MSC细胞/皿时,可生成不同比例的纯的CFUf集落,且数量与种入的细胞数呈良好的线性关系;小型CFUf集落再克隆生成CFUf集落的能力较弱,而大型CFUf集落具有较高的再克隆生成CFUf集落的能力,再克隆生成CFUf集落率高达90%,而且1个目的集落可再克隆生成多个CFUf集落;不同诱导方案均可诱导MSC生成脂肪细胞,而合用地塞米松(DM)、3异丁基1甲基黄嘌呤(IBMX)、胰岛素(IS)和消炎痛(ID)时效果最佳,诱导MSC生成脂肪细胞的效率可高达96%以上,且生成的细胞绝大多数为成熟脂肪细胞。结论采取贴壁细胞分离法经过长期传代可得到纯化的小鼠骨髓MSCs,MSCs具有高度增殖、自我更新和定向分化为脂肪细胞的能力;DM、IBMX、IS和ID4种诱导剂合用可诱导96%以上的该细胞定向分化为成熟脂肪细胞。

关键词: 小鼠, 间充质干细胞, 脂肪细胞, 组织工程

Abstract: ObjectiveTo establish separate and appraisal methods of murine bone marrow mesenchymal stem cells (MSCs) and optimize the suitable conditions inducting MSCs directional differentiating into adipocytes in vitro.MethodsThe differential adherence to plastic was employed to separate MSCs. CFU-f and successive CFU-f cultures were employed to characterize the potent of proliferation and self-renewal of MSCs. The different adipogenic medium was used as induction for the differentiation of MSCs into adipocytes. The differentiated cells were identified by oil red O immunohistochemistry stain.ResultsThe purified MSCs showed the morphology of fibroblasts. It was found that the number of CFU-f formation depended on the planted number of MSCs. It showed a good relationship. Small type colony of CFU-f had little potent to re-clone, but almost 90% big type colony of CFU-f had the potent to regenerate CFU-f. The MSCs could directionally differentiated into adipocytes induced by different adipogenic medium. But more than 96% MSCs differentiated into mature adipocytes when induced by combined with dexamethasone (DM), 1-methy-3-isobutylxanthine (IBMX), insulin (IS) and indomethacin (ID).ConclusionThe purified MSCs can be harvested by method of differential adherence to plastic, and these MSCs have the potent of proliferation and self-renewal. Moreover, more than 96% MSCs can differentiate into mature adipocytes when induced by combined with DM, IBMX, IS and ID.

Key words: murine, mesenchymal stem cells, adipocyte, tissue engineering