摘要
目的：研究SonicHedgehog（Shh）在雷奈酸锶（strontiumranelate，Sr）促进大鼠骨髓间充质干细胞（BMSCs）向成骨细胞分化中的作用。方法：采用全骨髓贴壁培养法分离、纯化、培养大鼠BMSCs，取第3-5代BM．SCs加入成骨诱导液诱导成骨分化，再加入不同浓度sr及Shh拮抗剂cyclopamine（Cy），分别观察它们对BMSCs向成骨细胞分化的影响。酶标法检测成骨细胞分化早期标志物碱性磷酸酶（alkalinephosphatase，ALP）的活性；茜素红染色检测细胞钙化水平；Westernblotting法检测Shh和Runx2蛋白的表达情况。结果：Sr（3mmol／L）可以使细胞ALP活性增高，钙结节形成增加。Sr（0．1-5mmol／L）作用BMSCs7d，可明显促进Shh和Runx2蛋白的表达，且Shh蛋白在1mmol／LSr作用时表达最多，而Runx2在3mmol／LSr作用时表达最多。1mmol／LSr作用BMSCs不同时间（1、3、5、7d），呈时间依赖性地上调Shh和Runx2蛋白的表达。cy（10Iμmol／L）不仅拮抗sr对Shh和Runx2表达的上调作用，还抑制sr对ALP和钙结节形成的促进作用。结论：sr可通过上调Shh蛋白及成骨特异性转录因子Runx2的表达促进BMSCs向成骨细胞分化。
AIM： To study the role of Sonic Hedgehog （Shh） in strontium ranelate （Sr）-induced osteogenic differentiation of rat bone marrow mesenchymal stem cells （BMSCs）. METI-IODS ： BMSCs was isolated from 4-week-old rats by adherent culture. The cells in the 3rd - 5th generations were induced to differentiate into obteoblasts, and then were treated with different concentrations of Sr and cyclopamine （Cy）. The activity of alkaline phosphatase （ALP） was mea- sured by colorimetry. Mineralized nodules were observed by alizarin red staining. The cellular Shh and Runx2 expression was detected by Western blotting. RESULTS ： Sr at concentration of 3 mmol/L increased the activity of ALP and induced the formation of mineralized nodules. Sr at concentrations ranging from 0.1 to 5 mmol/L increased the expression of Shh and Runx2 in the BMSCs at 7 d. Furthermore, the peak expression of Shh occurred following the exposure of Sr （ 1 mmol/ L） or Runx2 （3 mmol/L）. On the other hand, Sr at concentration of 1 mmoL/L showed a time-dependent increase in the expression of Shh and Runx2 from 1 d to 7 d. Cy at concentration of 10 ~mmol/L not only obviously inhibited Sr-induced expression of Shh and Runx2, but also antagonized the increase in the ALP activity and mineralization induced by Sr in the BMSCs. CONCLUSION： Sr promotes osteogenic differentiation of BMSCs by increasing the expression of Shh and Runx2.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2014年第1期159-164,共6页
Chinese Journal of Pathophysiology
基金
广东省自然科学基金资助项目(No.S2012010009403)
广东省科技计划(No.2011B031800002)
广州市科技计划(No.2012J4300082)
