Chinese Journal of Tissue Engineering Research ›› 2010, Vol. 14 ›› Issue (23): 4195-4198.doi: 10.3969/j.issn.1673-8225.2010.23.004

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In vitro proliferation and multi-directional differentiation of rat bone marrow mesenchymal stem cells

Xu Yong-tao1, Li Feng2, Lu Hou-geng1   

  1. 1Department of Orthopaedics, Jingzhou Hospital Affiliated to Tongji Medical College, Huazhong University of Science and Technology, Jingzhou   434020, Hubei Province, China;
    2Department of Orthopaedics, Tongji Hospital Affiliated to Tongji Medical College, Huazhong University of Science and Technology, Wuhan   430000, Hubei Province, China
  • Online:2010-06-04 Published:2010-06-04
  • About author:Xu Yong-tao, Doctor, Associate chief physician, Department of Orthopaedics, Jingzhou Hospital Affiliated to Tongji Medical College, Huazhong University of Science and Technology, Jingzhou 434020, Hubei Province, China xyt7512@medmail. com.cn

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Abstract:

BACKGROUND: Proliferation of bone marrow mesenchymal stem cells is not clear under some physiological condition; however, mitosis occurs following stimulation to a certain degree. The bone marrow mesenchymal stem cells have a strong proliferation ability and multi-directional differentiation potency.
OBJECTIVE: To further investigate the in vitro proliferation and multi-directional differentiation of bone marrow mesenchymal stem cells.
METHODS: Bone marrow mesenchymal stem cells were separated using whole bone marrow culture method and purified using attachment method. Morphology and growth characteristics were observed under inverted phase contrast microscope and growth curve was drawn using MTT method. Immunohistochemical method was used to identify cell surface markers CD44 of stem cells. The fourth-passage cells were respectively cultured in osteogenic, adipogenic and neuro-differentiation induction medium, and confirmed using the alkaline phosphatase staining, type II collagen immunohistochemical staining, oil red O staining and NeuN antibody immunohistochemical staining.
RESULTS AND CONCLUSION: After isolation and culture, cells developed a spindle or polygonal appearance. Growth curve was S-shaped, and population doubling time was 31 hours. Immunohistochemical staining demonstrated that CD44 expression was positive, but CD34 was negative. Two weeks after induction, calcium salt mineralization was found in the bone marrow mesenchymal stem cells of passage 4. Two weeks after chondrogenic induction, type II collagen demonstrated a positive expression. Two weeks after fat-induced culture, a great quantity of red fat drops were observed in cytoplasm. Six hours after neural induction, cells displayed processes, which were similar to axon and dendrite fiber. NeuN immunohistochemical staining demonstrated a positive result. This suggested that in vitro cultured bone marrow mesenchymal stem cells had a strong proliferation and could differentiate into osteoblasts, chondrocytes, adipocytes, and neuronal-like cells.

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