BMP-2文章--插图版final.docVIP

BMP-2对大鼠骨髓间充质干细胞成骨作用的影响 孙健 余优成 顾章愉 顾亮 毕玮 【摘要】 目的　通过携带骨形态发生蛋白-2（bone morphogenetic protein-2; BMP-2）基因的慢病毒载体感染大鼠骨髓间充质干细胞(bone mesenchym alstem cells, BMSCs)后，观察BMP-2对BMSCs成骨作用的影响。方法　原代培养大鼠BMSCs并进行传代及鉴定，用携带BMP-2的慢病毒感染第三代BMSCs后，进行骨向诱导，通过茜素红染色观察钙沉积Ⅰ及smad表达的影响。 结果　①原代培养出大鼠骨髓间充质干细胞并进行鉴定②携带BMP-2和EGFP的慢病毒感染第三代BMSCs后，免疫荧光检测和RT-PCR及WB检测感染成功③大鼠骨髓间充质干细胞感染慢病毒后进行骨向诱导，钙沉积明显增加Ⅰ及smad表达增加。结论　BMP-2可明显促进大鼠骨髓间充质干细胞向成骨细胞定向分化，为BMSCs作为组织工程研究的种子细胞提供实验支持。2；大鼠骨髓间充质干细胞；成骨诱导　 The study of BMP-2 in the osteogenic effect of rat mesenchymal stem cell SUN Jian,YU You-cheng,GU Zhang-yu,GU Liang,BI Wei Department of Stomatology,Zhongshan Hospital, Fudan University , Shanghai 200032 China Corresponding author:YU You-cheng,Email:yuyoucheng@ 【Abstract】　　　To observe th role of BMP-2 in the osteogenic effect of rat mesenchymal stem cell isolate (BMSCs). 　Methods 　The primary culture of rat BMSCs was succeeded and then was identified. The rat BMSCs were infected with the recombinant lentivirus with BMP-2. The osteogenic effect with BMP-2 was observed. In addition, adhesive ability of BMP-2-BMSCs was detected through adhesion assay and expression of osteogenic factors OPN、OCN、Col-Ⅰ and smad was observed by RT-PCR and western blot. Results Rat BMSCs were cultured and identified successfully. The osteogenic effect of BMSCs was improved by BMP-2. Lenti-BMP-2 BMSCs adhesive potential enhanced and osteogenic factors were up-regulated.Conclusions BMP-2 may facilitate the osteogenic effect of the rat BMSCs and would provide favourable cell resource for tissue engineering. 【Key words】bone morphogenetic protein 2mesenchymal stem cell; osteogenic effect 近年来，随着骨组织工程研究的不断深入,骨髓间充质干细胞(bone mesenchym alstem cells, BMSCs)日益受到关注。 因其具有多向分化潜能而逐渐成为骨组织工程中首选的种子细胞[1]。本实验拟将骨形态发生蛋白（bone morphogenetic protein）BMP -2通过慢病毒载体感染入BMSCs，观察BMP-2对骨髓间充质干细胞骨向分化能力的影响。 材料和方法 1. 材料： （1）动物：取8周龄Sprague-Dawley 大鼠1只，体重180克， 雌雄不限（复旦大学上海医学院实验动物中心提供）。 （2）病毒载体构建：携带BMP-2 和增强型绿色荧光蛋白（enhanced green fluorescent protein, EGFP)基因的慢病毒表达载体，由上海吉凯公司协助构建， 通过扩增HT