Chinese Journal of Tissue Engineering Research ›› 2012, Vol. 16 ›› Issue (10): 1742-1747.doi: 10.3969/j.issn.1673-8225.2012.10.008

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Single-cell clonal culture and identification of human bone marrow stromal stem cells  

Teng Yong1, Xu Jian-li2, Li Xu-sheng1, Hu Yun-yu3, Wang Zhen3   

  1. 1First Department of Orthopedics, Urumqi General Hospital of Lanzhou Command, PLA, Urumqi 830000, Xinjiang Uygur Autonomous Region, China; 2Department of Hematology, First Affiliated Hospital of Xinjiang Medical University, Urumqi  830000, Xinjiang Uygur Autonomous Region, China;      3Institute of Orthopedics, Xijing Hospital, Fourth Military Medical University of Chinese PLA, Xi’an  710032, Shaanxi Province, China
  • Received:2011-09-24 Revised:2012-02-01 Online:2012-03-04 Published:2012-03-04
  • About author:Teng Yong☆, M.D., Associate chief physician, Master’s supervisor, First Department of Orthopedics, Urumqi General Hospital of Lanzhou Command, PLA, Urumqi 830000, Xinjiang Uygur Autonomous Region, China orthtengyong@163.com
  • Supported by:

    Medical Science and Research Development of Lanzhou Command, No.LXH2007014*; the National Natural Science Foundation of China, No.51165044*; a grant from the Twelfth “Five-Year” Development Program Foundation of Chinese PLA, No.CWS11J01*

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Abstract:

BACKGROUND: Bone marrow stromal stem cells lack of specific surface marker and their identification challenges scholars all the time.
OBJECTIVE: To explore the culture condition for clonal isolation of human bone marrow stromal stem cells (BMSCs) and identify their surface markers and differentiation potentials.
METHODS: Human bone marrow was taken and BMSCs were isolated using density gradient centrifugation. Clone-like cells were selected by single cell limiting dilution. The surface antigens of the cloned BMSCs were analyzed by flow cytometry and immunocytochemistry. Multi-differentiation capacities were evaluated by inducing BMSCs into osteoblasts, chondrocytes and cadiocytes. RT-PCR and immunocytochemistry were applied to detect the three mutli-differentiations.
RESULTS AND CONCLUSION: Single cell-derived BMSCs could be expanded to passage 28 in vitro which still maintaued active proliferation ability.The expanded BMSCs expressed CD29, CD44, CD106, but not CD14, CD34, CD45, HLA-DR. They could be induced to differentiate into osteoblasts, chondrocytes and cadiocytes. These findings suggest that cloned BMSCs can maintain the phenotypes of stem cells during in vitro culture.

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