Chinese Journal of Tissue Engineering Research ›› 2010, Vol. 14 ›› Issue (2): 262-266.doi: 10.3969/j.issn.1673-8225.2010.02.017

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Effect of eucommia bark on osteopontin and osteoprotegerin expressions in rat bone marrow stem cells: Is there any difference between water extract and methanol extract?

Zhang Yan-hong1, Xie Huan-song1, Xia Shu-lin2, Zhao Chun2, Tan Xiang-ling2   

  1. 1 Department of Cell Biology, Life School of Nantong University, Nantong  226007, Jiangsu Province, China; 2 Institute of Bioengineering, Nantong University, Nantong  226007, Jiangsu Province, China
  • Online:2010-01-08 Published:2010-01-08
  • Contact: Tan Xiang-ling, Master, Professor, Institute of Bioengineering, Nantong University, Nantong 226007, Jiangsu Province, China tanxl@ntu.edu.cn
  • About author:Zhang Yan-hong, Associate professor, Department of Cell Biology, Life School of Nantong University, Nantong 226007, Jiangsu Province, China ntuzyh@126.com
  • Supported by:

    the Social Development Foundation of Wuxi, No. CSE00713*

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Abstract:

BACKGROUND: Previous studies demonstrated that eucommia bark can promote bone marrow stem cells (BMSCs) differentiated into osteoblasts, but relative mechanism is poorly understood. 
OBJECTIVE: To investigate the effects of eucommia bark water/methanol extracts on expressions of osteopontin (OPN) and osteoprotegerin (OPG) in rat BMSCs.
METHODS: Totally 2 g eucommia bark powder were added into water or methanol to 15 mL and oscillated for 1 hour at room temperature. After soaked overnight, both extracts were centrifuged at 15 000 r/min for 10 minutes. Water extract was obtained from supernatant in water soaked powder. In methanol soaked powder, methanol extracts was obtained by concentrated supernatant in vacuo and resolved using 15 mL water. Water and methanol extracts were then filtered by 0.22 μm membrane, and conserved at -20 ℃. Six SD rats, aged 2 months, were selected, and the 3rd passage of BMSCs were induced by water or methanol extracts with dilution of 1×10-2, 1×10-3, 1×10-4 and 1×10-5, respectively. PBS was added in the negative control group. All cells were cultured for 6 days. Expressions of OPN and OPG was measured by immunocytochemistry at 6 days with induction. The expression of OPN and OPG induced by water and methanol at 1×10-3 and 1×10-4 dilution was detected by RT-PCR.
RESULTS AND CONCLUSION: Immunocytochemistrical results indicated that both water and methanol extracts of eucommia bark simulated OPN and OPG expression, in particular with dilution of 1×10-4. The methanol extracts had a stronger effect than water extract, but the expression of OPG did not change obviously. RT-PCR demonstrated that at the 3rd day of inducement, the level of OPN expression induced by water extract was higher than that of methanol extract, and no OPG expression was detected. Osteogenic differentiation of rat MSCs induced by eucommia bark water/methanol extracts relates to stimulating expression of OPN, which has no correlation to OPG. OPN expression induced by water extract is early than that of methanol extract. 

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