中国组织工程研究

中国组织工程研究 ›› 2012, Vol. 16 ›› Issue (1): 27-30.doi: 10.3969/j.issn.1673-8225.2012.01.005

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

全骨髓直接贴壁分离人骨髓间充质干细胞的生物学特性****☆

张  颢1，2，张  斌2，程  梅1，陶艳玲1，扈江伟2，徐  曼2，陈  虎2    

  1. 1哈尔滨市第一医院哈尔滨血液病肿瘤研究所，黑龙江省哈尔滨市  150010；2解放军军事医学科学院附属医院全军造血干细胞移植中心，北京市100071 哈尔滨市第一医院和解放军军事医学科学院附属医院为共同第一完成单位。
  • 收稿日期:2011-10-25 修回日期:2011-11-24 出版日期:2012-01-01 发布日期:2012-01-01
  • 通讯作者: 通讯作者：陈虎，博士，主任医师，解放军军事医学科学院附属医院全军造血干细胞移植中心，北京市 100071 chenhu217@ yahoo.com.cn
  • 作者简介:张颢☆，男，1973年生，河南省息县人，汉族，2008年解放军福建医科大学毕业，博士，副主任医师，主要从事干细胞基础与临床应用的研究。gx-zhanghao@126.com 并列第一作者：张斌，男，1971年生，河南省郑州市人，汉族，2002年解放军第四军医大学毕业，博士，副主任医师，主要从事干细胞及生物治疗基础与临床应用方面的研究。zb307ctc@163.com
  • 基金资助:

    哈尔滨市科技创新人才研究专项基金项目(2009RFQQS027)，课题名称：胎盘间充质干细胞规模化扩增及移植中应用；黑龙江省卫生厅科研课题(2011-518)，课题名称：脐带间充质干细胞治疗移植物抗宿主病的临床研究；国家863重大专项课题(2011AA020114)，课题名称：干细胞应用于恶性血液病放化疗后造血损伤再生修复的研究及临床评价；首都临床特色应用研究(SQ2010 AA0201008009)，课题名称：脐带间充质干细胞治疗移植物抗宿主病。

Isolation and charcaterization of human bone marrow mesenchymal stem cells using a whole bone marrow adherent culture technique

Zhang Hao1, 2, Zhang Bin2, Cheng Mei1, Tao Yan-ling1, Hu Jiang-wei2, Xu Man2, Chen Hu2   

  1. 1Harbin Institute of Hematology Oncology, the First Hospital of Harbin, Harbin  150010, Heilongjiang Province, China; 2Hematopoietic Stem Cell Transplantation Center, Affiliated Hospital of Academy of Military Medical Science of Chinese PLA, Beijing  100071, China The First Hospital of Harbin and the Affiliated Hospital of Academy of Military Medical Science of Chinese PLA were co-considered as the first units.
  • Received:2011-10-25 Revised:2011-11-24 Online:2012-01-01 Published:2012-01-01
  • Contact: Correspondence to: Chen Hu, Doctor, Chief physician, Hematopoietic Stem Cell Transplantation Center, Affiliated Hospital of Academy of Military Medical Science of Chinese PLA, Beijing 100071, Chinachenhu217@yahoo.com.cn
  • About author:Zhang Hao☆, Doctor, Associate chief physician, Harbin Institute of Hematology Oncology, the First Hospital of Harbin, Harbin 150010, Heilongjiang Province, China; Hematopoietic Stem Cell Transplantation Center, Affiliated Hospital of Academy of Military Medical Science of Chinese PLA, Beijing 100071, Chinagx-zhanghao@126.com Zhang Bin☆, Doctor, Associate chief physician, Hematopoietic Stem Cell Transplantation Center, Affiliated Hospital of Academy of Military Medical Science of Chinese PLA, Beijing 100071, Chinazb307ctc@163.com

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摘要：

背景：体外分离培养骨髓间充质干细胞成为实验研究和临床应用的关键环节。
目的：采用全骨髓培养法分离人骨髓间充质干细胞，建立一种简单、有效的分离培养骨髓间充质干细胞的方法。
方法：通过全骨髓培养法分离培养人骨髓间充质干细胞，并通过传代进行纯化和扩增培养。分别在特定诱导体系中诱导人骨髓间充质干细胞向脂肪、成骨及软骨细胞分化。
结果与结论：采用全骨髓培养分离法能获得90%以上纯度的骨髓间充质干细胞；流式细胞仪检测结果显示，贴壁细胞均表达CD73、CD90、CD105，不表达造血细胞表型CD34、CD45和HLA-DR；细胞倍增时间为(24.04±0.49) h；细胞周期分析表明：G0~G1期和S+G2+M期所占比例分别为71.63%和28.37%；诱导分化结果显示，人骨髓间充质干细胞能够向脂肪、骨和软骨细胞分化。说明全骨髓分离培养法是一种较好的分离人骨髓间充质干细胞的方法。

关键词: 全骨髓培养, 间充质干细胞, 分离, 培养, 分化

Abstract:

Abstract
BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) are widely used in cell therapy and tissue regeneration, so their isolation and culture are a key technique for research and clinical application.
OBJECTIVE: To isolate the BMSCs by whole bone marrow adherent culture technique and to establish an easy and effective method to isolate and culture BMSCs.
METHODS: The BMSCs were isolated by whole bone marrow adherent culture technique, and purified and amplified by passage. BMSCs were induced to differentiate into adipocytes, osteocytes and chondrocytes in special differentiation condition.   
RESULTS AND CONCLUTION: The purity of the BMSCs isolated by whole bone marrow adherent culture technique was more than 90%. The adherent cells displayed an abundant presence of CD73, CD90, CD105 and absence of Hematopoietic cell phenotype CD34, CD45 and HLA-DR which were detected by flow cytometry. The cell doubling time was (24.04±0.49) hours; Cell cycle showed that there was percentage of stem cells as G0 /G1 71.63% and S+G2+M 28.37% respectively. The induced and differentiated results showed than BMSCs could be differentiated into adipocytes, osteocytes and chondrocytes. Based on these findings, the whole bone marrow adherent culture technique is a better method to obtain MSCs from human bone marrow.

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