Chinese Journal of Tissue Engineering Research ›› 2012, Vol. 16 ›› Issue (19): 3421-3426.doi: 10.3969/j.issn.1673-8225.2012.19.001

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Effects of silencing annexin A1 gene on proliferation and osteogenic differentiation of rabbit bone marrow mesenchymal stem cells 

Liang Bo-wei1, Pan Xin-yuan2, Zhao Jin-min1, Yin Guo-qian2, Hu Feng1   

  1. 1Department of Traumatic Orthopedics and Hand Surgery, 2Department of Plastic and Cosmetic Surgery, First Affiliated Hospital of Guangxi Medical  University, Nanning 530021, Guangxi Zhuang Autonomous Region, China
  • Received:2012-02-06 Revised:2012-03-15 Online:2012-05-06 Published:2012-05-06
  • Contact: Zhao Jin-min, M.D., Doctoral supervisor, Professor, Department of Traumatic Orthopedics and Hand Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China zhaojinmin@126.com
  • About author:Liang Bo-we☆, Studying for doctorate, Attending physician, Department of Traumatic Orthopedics and Hand Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China LBW5515@163.com Pan Xin-yuan☆, Studying for master’s degree, Department of Plastic and Cosmetic Surgery, First Affiliated Hospital of Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, China u2coldplay@163.com
  • Supported by:

    the National Natural Science Foundation of China, No. 30860285*

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Abstract:

BACKGROUND: Annexin A1 (ANXA1) protein is involved in regulation of cell proliferation and differentiation. Steroid-induced osteonecrosis of femoral head is related to altered proliferation and osteogeneic differentiation of bone marrow mesenchymal stem cells (BMSCs).
OBJECTIVE: To investigate the effects of silencing ANXA1 gene on proliferation and osteogenic differentiation of rabbit BMSCs.
METHODS: New Zealand white rabbit BMSCs were cultured in vitro and divided into three groups. In the RNA interference group, BMSCs were transfected with lentiviral vectors carrying short hairpin RNA (LV-shANXA1) to down regulate the expression of ANXA1 gene. In the negative control group, BMSCs were transfected with lentiviral vectors carrying nonsense gene sequences (LV-shANXA1-NC). In the blank control group, there was no intervention.
RESULTS AND CONCLUSION: The positive rate of CD44, CD105 and CD45 in BMSCs after purification was 97.2%, 95.8 % and 2.5%, respectively. The infection efficiency of lentivirus was 80% at multiplicity of infection (MOI) of 50 and 95% at MOI of 100 in BMSCs after selection by puromycin. Real-time PCR and western blot results showed that the silencing efficiency was above 72.4% at MOI of 50. After down regulating the expression of ANXA1 gene, significant growth inhibition could be found in BMSCs; the alkaline phosphatase positive rate and alkaline phosphatase activity decreased, and the alizarin red staining results showed negative reaction in the subsequent induction of osteogenic differentiation in the cells. The experimental results show that silencing ANXA1 gene reduces the proliferation and osteogenic differentiation capacity of BMSCs, which may be one of mechanisms underlying steroid-induced avascular necrosis of femoral head.

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