中国组织工程研究

中国组织工程研究 ›› 2011, Vol. 15 ›› Issue (1): 12-16.

• 骨髓干细胞 bone marrow stem cells • 上一篇    下一篇

低渗结合自然沉降法分离兔骨髓间充质干细胞及其鉴定

林春博1，杨  渊1，陈维平2   

  1. 1广西骨伤医院，广西壮族自治区南宁市 530012
    2广西医科大学，广西壮族自治区南宁市  530021
  • 收稿日期:2010-07-25 修回日期:2010-10-14 出版日期:2011-01-01 发布日期:2011-01-01
  • 通讯作者: 杨渊，教授，广西骨伤医院，广西壮族自治区南宁市 530012; 陈维平，教授，广西医科大学，广西壮族自治区南宁市 530021
  • 作者简介:林春博★，男，1983年生，广西壮族自治区玉林市人，汉族，2010年广西中医学院毕业，硕士，医师，主要从事骨关节创伤及脊柱疾病的中医药防治研究。 linchunbo1983@163.com
  • 基金资助:

    广西科技厅资助项目(桂科攻0816004-15)，广西科技厅资助项目(桂科自0728077)，广西科技厅资助项目(桂科自0832157)，广西科技厅资助项目(桂科攻0592007-1G)。

Isolation and identification of rabbit bone marrow mesenchymal stem cells by low permeability combined with natural sedimentation

Lin Chun-bo1, Yang Yuan1, Chen Wei-ping2   

  1. 1Guangxi Bone Trauma Hospital, Nanning  530012, Guangxi Zhuang Autonomous Region, China
    2Guangxi Medical University, Nanning  530021, Guangxi Zhuang Autonomous Region, China
  • Received:2010-07-25 Revised:2010-10-14 Online:2011-01-01 Published:2011-01-01
  • Contact: Yang Yuan, Professor, Guangxi Bone Trauma Hospital, Nanning 530012, Guangxi Zhuang Autonomous Region, China; Chen Wei-ping, Professor, Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China
  • About author:Lin Chun-bo★, Master, Physician, Guangxi Bone Trauma Hospital, Nanning 530012, Guangxi Zhuang Autonomous Region, China linchunbo1983@163.com
  • Supported by:

    Grants of Department of Science and Technology of Guangxi Zhuang Autonomous Region of China, No. Guikegong 0816004-15; Guikezi 0728077; Guikezi 0832157; Guikegong 0592007-1G

PDF

679

被引次数

16

摘要：

背景：骨髓间充质干细胞数量极少，10万个骨髓有核细胞中才有1个骨髓间充质干细胞，如何成功分离并鉴定骨髓间充质干细胞是研究中的关键工作。
目的：建立完善的骨髓间充质干细胞体外分离培养体系，从细胞的形态、表型和功能方面鉴定骨髓间充质干细胞。
方法：低渗原理去除杂细胞，结合自然沉降分离骨髓间充质细胞，并对其形态学特征进行光镜观察。对骨髓间充质细胞的表型进行流式细胞仪检测。并将骨髓间充质细胞向成骨细胞和脂肪细胞进行诱导分化，行形态学观察和碱性磷酸酶、Von kossa、Ⅰ型胶原、油红O染色检测。
结果与结论：实验分离获取的骨髓间充质细胞经表型鉴定为：CD29(+)，CD45(-)；向成骨细胞诱导后，碱性磷酸酶、矿化结节染色及Ⅰ型胶原检测均为阳性；向脂肪细胞诱导后，油红O染色阳性。结果表明实验分离的骨髓间充质细胞为类成纤维细胞样非造血类干细胞，此类细胞具有向成骨细胞和脂肪细胞分化的功能。

关键词: 骨髓间充质干细胞, 分离, 鉴定, 低渗原理, 自然沉降, 细胞形态, 细胞表型, 细胞功能

Abstract:

BACKGROUND: The number of bone marrow mesenchymal stem cells (BMSCs) was few. 100 000 bone marrow karyocytes contain 1 BMSC. How to successfully isolate and identify BMSCs is a key in the study.
OBJECTIVE: To establish a perfect culture system in vitro of BMSCs; BMSCs were identified from cell morphology, phenotype and functional aspects.
METHODS: Low permeability principle was used to remove mixed cell, and combined with the natural sedimentation to isolate BMSCs. Their morphological characteristics were observed by light microscopy. The cell phenotype of BMSCs was determined by flow cytometry. BMSCs were induced into osteoblasts and adipocytes. Cell morphology was observed and alkaline phosphatase (AKP), Von Kossa, collagen typeⅠ, oil red O detection were performed.
RESULTS AND CONCLUSION: BMSCs were identified as phenotype: CD29(+), CD45(-). Following being induced into osteoblasts, AKP, mineralized nodules and collagen type Ⅰ staining were detected positively; to adipocytes, the oil red O staining was detected positively. Results suggest that isolated BMSCs are non-hematopoietic stem cells with fibroblast-like cell morphology. These cells had the functions of differentiating into osteoblasts and adipocytes.

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