Chinese Journal of Tissue Engineering Research ›› 2010, Vol. 14 ›› Issue (45): 8365-8368.doi: 10.3969/j.issn.1673-8225.2010.45.002

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Optimization of small interfering RNA-mediated RhoA silencing in bone marrow mesenchymal stem cell cultures

Lü Gang1, Yao Xin2   

  1. 1 Postgraduate College, Tianjin Medical University, Tianjin  300000, China; 2 Department of Neurosurgery, Huanhu Hospital of Tianjin, Tianjin  300000, China
  • Online:2010-11-05 Published:2010-11-05
  • Contact: Yao Xin, Master, Chief physician, Master’s supervisor, Department of Neurosurgery, Huanhu Hospital of Tianjin, Tianjin 300000, China phoenix6a6@163.com
  • About author:Lü Gang★, Studying for master’s degree, Attending physician, Postgraduate College, Tianjin Medical University, Tianjin 300000, China lgemail@eyou.com

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Abstract:

BACKGROUND: Previous culture method of bone marrow mesenchymal stem cells (BMSCs) has problems such as senescence and low differentiation rate.
OBJECTIVE: To determine whether the RhoA gene silencing can optimize BMSC cultures.
METHODS: BMSCs from Sprague Dawley rats were cultured in vitro, and were transfected by small interfering RNA (siRNA) to knock down the expression of RhoA. BMSCs were divided into three groups: stem cell group (non-transfection of siRNA), disrupted by random sequence siRNA transfection stem cell group, and siRNA transfection of stem cell group. Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot assay were used to assess RhoA gene and protein expression before and after transfection. Cellular proliferation optimization was determined by the cell growth curve and MTT assay. The cell cycle was analyzed by flow cytometry.
RESULTS AND CONCLUSION: Expression of RhoA gene and protein was significantly reduced following transfection of siRNA compared with stem cell group (P < 0.05). The cell growth speed was significantly increased, the cell cycle of G0/G1 phase was decreased and cell number in S phase was increased (P < 0.05). These indicated that the RhoA gene silencing can promote BMSC proliferation, and optimize culture methods. 

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