中文摘要：

目的：探讨蛇床子素对体外培养的大鼠成骨细胞增殖的影响及机制。方法：新生SD大鼠10只，脱颈处死后取出颅盖骨，剪成约1mm×lmm的碎片，采用胰酶预消化15min，继以0．1％I型胶原酶37℃消化2次，每次1h，所得细胞混匀后置于5％C02培养箱中37℃恒温培养。细胞培养48h、28d后行AIJP染色和矿化结节染色以进行成骨性鉴定。细胞随机分为5组，分别加入终浓度为100、50、25、12．5、0μmol／L的蛇床子素进行干预。24、48、72h后采用CCK-8法检测细胞增殖率，并通过WestemBlot方法检测加药1周后PCNA、β-catenin蛋白表达量。结果：镜下观察细胞形态不规则，具有典型的成骨细胞形态；ALP染色及矿化结节染色均为阳性。CCK-8检测发现100txmol／L的蛇床子素干预24h后即可抑制成骨细胞生长，50μmol／L和25μmol／L的蛇床子素于加药后48h开始抑制成骨细胞生长，12．5μmol／L蛇床子素对成骨细胞的增殖无显著影响。Western Blot检测结果显示PCNA及B-catenin随蛇床子素浓度增加而表达下调。结论：100、50、25μmol／L各个浓度的蛇床子素可抑制成骨细胞增殖，12．5μmol／L蛇床子素对细胞的增殖特性无显著影响。蛇床子素抑制成骨细胞增殖的作用可能是通过干扰Wnt／β-catenin信号通路而实现的。

英文摘要：

Objective：To investigate the effects of osthole on proliferation of neonatal rat osteoblast and the mechanism. Methods ： Ten 24 hours old SD rats were executed by dislocating. The cranium of rats were removed and cut into blocks of 1 mm×l mm size. After digested by trypsin for 15 min ,the cranium were digested by type I collagenase for one hour two times. The mixed cells were cultured in thermostat incubator with 5% CO2 under the condition of 37℃. To identify the cells, ALP staining and alizarin red staining were performed after cultured 48 h and 28 d. The osteoblasts were randomly divided into five groups. Ceils were treated with osthole at concentrations of 100,50,25,12.5,0 μmol/L. CCK-8 method was used to evaluate the proliferation after 24 h, 48 h and 72 h. The expression of PCNA and 13-catenin protein were detected through the method of Western Blot after one week. Results ：The cells had irregular shapes and showed typical features of osteoblast. The results of ALP staining and alizarin red staining were both positive. CCK-8 detection showed that the osthole with final concentration of 100 p, mol/L inhibited the proliferation of osteoblast after 24 h,while the osthole with final concentrations of 50 μmol/L and 25 μmol/L displayed the inhibition effect after 48 h. The osthole of 12.5μmol/L had no obvious influence on the proliferation of osteoblast. The result of Western Blot showed that osthole reduced the expression of PCNA and β-catenin protein in a dose- dependent manner. Conclusion：The osthole with final concentrations of 100,50,25 μmol/L inhibited the proliferation of os- teoblast （P〈0.05）. The osthole with final concentrations of 12.5 μmol/L had no obvious influence on the proliferation of os- teoblast （P〉0.05）. These findings demonstrate that osthole may inhibit the proliferation of osteoblast by regulating the Wnt/β- catenin signaling in osteoblast.