淫羊藿苷与染料木黄酮对体外培养成骨细胞增殖及矿化成熟影响的对比研究  

Comparative study on effect of icariin and genistein on proliferation and mineralization of osteoblasts in vitro

作　　者：明磊国[1] 陈克明[1] 葛宝丰[1] 马慧萍[2] 周建[1] 

机构地区：[1]兰州军区兰州总医院骨研所,甘肃兰州730050 [2]兰州军区兰州总医院药材科,甘肃兰州730050

出　　处：《中国中药杂志》2011年第16期2240-2245,共6页China Journal of Chinese Materia Medica

基　　金：甘肃省科技重大专项计划项目(092nkDA025)

摘　　要：目的:对比研究淫羊藿苷(icariin,I)与染料木黄酮(genistein,G)对体外培养大鼠成骨细胞(rat calvarial osteo-blasts,ROB)增殖与分化成熟的影响及其药理活性的强弱。方法:取新生SD大鼠颅骨,多次酶消化法获得成骨细胞,培养于含10%FBS的MEM培养液中,3 d后首次换液,铺满90%皿底后传代培养;I与G的作用终浓度为1×10-5 mol.L-1,增殖分析采用96孔板,MTT法分析;分化成熟分析采用24孔板,于成骨性诱导培养第3,6,9,12天分别测碱性磷酸酶(alkaline phospha-tase,ALP)活性、钙盐沉积量、骨钙素,第12天进行ALP组织化学染色、茜素红染色和钙化结节计数;于成骨性诱导培养0,6,12,24,48,72 h提取总RNA,Real-time RT-PCR法检测Runx-2,bFGF,IGF-Ⅰ,Osterix(OSX)等成骨性相关因子的表达情况。结果:终浓度为1×10-5 mol.L-1的I与G均不促进ROB的增殖,均能显著提高细胞ALP活性、增加钙含量、促进骨钙素分泌,并增加钙化结节数量;同时上调Runx-2,bFGF,IGF-Ⅰ和Osterix等成骨性相关因子的表达量;在成骨细胞水平上I促进ROB矿化成熟的药理学活性明显强于G的活性。结论:1×10-5 mol.L-1的I与G均能促进ROB矿化成熟,均有抗骨质疏松的作用,但在成骨细胞水平上I促进ROB矿化成熟的药理学活性显著强于G。Objective： To investigated the effect of icariin and genistein on proliferation and mineralization of cultured rat osteoblasts（rat calvarial osteoblasts,ROB）.And to contrast the pharmacological activity of icariin and genistein.Method： Bone cells were obtained by enzyme digestion from the segregated neonatal SD rat skull,and were cultured in MEM containing 10% FBS which was changed after three days later.Serial subcultivation was proceeded when cells covered with 90% culture dish.The final action concentration of icariin and genistein were both 1×10-5 mol·L-1.Proliferation was analyzed by MTT on 96-well plates,while differentiation was analyzed on 24-well plates.Under the induced condition,the alkaline phosphatase activity,calcium salt sediment yield and osteocalcin were measured at the 3,6,9,12 d.At 12th day,ALP staining,alizarin red staining and calcified nodule count were preceded.Total RNA was isolated at 0,6,12,24,48,72 h.The gene expression of bFGF,IGF-1,Osterix and Runx-2 was analyzed by Real-time RT-PCR.Result： With the concentration of 1×10-5 mol·L-1,icariin and genistein have no significant effect on the ROB′s proliferation.The osteogenesis,ALP activity,calcium salt sediment yield and osteocalcin,calcified tubercle amount were significantly increased.And they enhanced the mRNA level of bFGF,IGF-1,Osterix and Runx-2.On the level of osteoblasts,the activity of icariin is stronger than that of genistein.Conclusion： When the final concentration of icariin and genistein is 1×10-5 mol·L-1,they can significantly promoted ROB maturation.And on the level of osteoblasts,the activity of icariin is stronger than that of genistein.

关 键 词：成骨细胞 淫羊藿苷 染料木黄酮 骨钙素 碱性磷酸酶 Runx-2 bFGF IGF-Ⅰ OSTERIX 

分 类 号：R285.5[医药卫生—中药学]