Chinese Journal of Tissue Engineering Research ›› 2012, Vol. 16 ›› Issue (14): 2515-2519.doi: 10.3969/j.issn.1673-8225.2012.14.009

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In vitro culture, identification and osteogenic differentiation of human bone marrow mesenchymal stem cells

Liu Wei1, Liu Meng2, Zhu Jing-song1, Hao Hong-wei1, Dong Ning-zheng2, Zhou Hai-bin1   

  1. 1Department of Orthopedics, Second Affiliated Hospital of Soochow University, Suzhou 215004, Jiangsu Province, China; 2Tangzhongying Research Center of Hematology, Soochow University, Suzhou  215004, Jiangsu Province, China
  • Received:2012-01-10 Revised:2012-01-28 Online:2012-04-01 Published:2012-04-01
  • Contact: 周海斌，博士，副主任医师，苏州大学附属第二医院骨科，江苏省苏州市 215004 zhouhaibins@yahoo.com.cn
  • About author:Liu Wei★, Studying for master’s degree, Department of Orthopedics, Second Affiliated Hospital of Soochow University, Suzhou 215004, Jiangsu Province, China aqqvvvv@126.com

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Abstract:

BACKGROUND: Research approaches and measurement index regarding osteogenic differentiation of bone marrow mesenchymal stem cells in vitro are not fully considered at home and abroad.
OBJECTIVE: To establish and consummate a set of methods of isolating, culturing and identifying human bone marrow-derived mesenchymal stem cells (hBMSCs) and investigate their osteogenic differentiation ability in vitro.
METHODS: hBMSCs were isolated using the method of density gradient centrifugation. The cell surface phenotypes of BMSCs were identified by flow cytometry. Passage 3 hBMSCs were induced by osteogenic culture medium for 2-3 weeks.
RESULTS AND CONCLUSION: The primary cultured hBMSCs grew and proliferated vigorously after subculture. Passage 3 hBMSCs were positive for CD44, CD73, CD90 and were negative for CD34. After osteogenic induction, alkaline phosphatase activity was increased and cells were stained positive by Gomori, Von kossa and alizarin red. RT-PCR results showed the gene expression of collagen Ⅰ, alkaline phosphatase, osteocalcin, bone sialoprotein, osteopotin and osteonectin. This confirms that the primary hBMSCs can differentiate into osteoblasts in vitro successfully. These findings suggest that a stable, mature method of isolating, culturing and amplifying hBMSCs has been established.

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