Cbfa1因子对成纤维细胞中软骨相关基因表达的影响  

Cbfa1′s effect on chondrogenic gene expression in fibroblast

作　　者：张伟[1] 朱雅萍[1] 王君[1] 邓廉夫[1] 

机构地区：[1]第二医科大学附属瑞金医院上海市伤骨科研究所,上海200025

出　　处：《中华医学杂志》2005年第16期1129-1132,共4页National Medical Journal of China

基　　金：国家自然科学基金 ( 30070759 );上海市青年科技启明星(H)计划资助项目(01QMH1409)

摘　　要：目的　研究转录因子Cbfa1在软骨发生中的作用。方法　应用逆转录聚合酶链反应酶技术,直接从E13 5d胎鼠肢芽细胞中克隆出Cbfa1全长cDNA,并将其定向克隆到pcDNA3 1质粒载体中,经酶切鉴定和序列分析证实了克隆的正确性。在构建出Cbfa1真核表达载体后,将其转化到成纤维细胞中,48h后比较转染组和未转染组Sox9、Sox5和Sox6基因的转录表达情况,72h后比较转染组和未转染组中Ⅱ型胶原的表达。结果　Cbfa1在成纤维细胞中的高表达能提高Sox9和Ⅱ型胶原的表达,但Cbfa1在成纤维细胞中的高表达对Sox5和Sox6的表达没有影响。结论　Cbfa1可能参与了软骨细胞的发育调控。Objective Core-binding factor a1, Cbfa1, which belongs to the runt-domain gene family, is an essential transcription factor for osteoblastic differentiation and osteogenesis. To examine the effects of Cbfa1 exhibit on gene expression which involving in the chondrogenesis. Methods On the first, using RT-PCR method, we directly cloned Cbfa1/Runx2 full length cDNA from E13.5 days mouse embryos limb buds. Consequently Cbfa1/Runx2 was cloned into pcDNA3.1 plasmid to make a eukaryotic expression vector. All the clones were proved to be correct by enzyme cutting and sequencing analysis. On the second, we used pcDNA3.1-Cbfa1 to tansfect fibroblast. 48 hours later, we contrasted the gene transcription of Sox9、Sox5 and Sox6 between tansfected and untransfected.72 hours later, type Ⅱcollagen have also been examined by Western bloting. Results Cbfa1 overexpression in fibroblast can upregulate the expression of Sox9 and type Ⅱcollagen, but it have no effect on the expression of Sox5 and Sox6. Conclusion Cbfa1 may involve in the regulation of chondrogenesis.

关 键 词：CBFA1 成纤维细胞 相关基因表达 逆转录聚合酶链反应 全长cDNA 真核表达载体 Sox5 Ⅱ型胶原 转录因子 方法应用 肢芽细胞 质粒载体 定向克隆 序列分析 酶切鉴定 表达情况 72h后 发育调控 软骨细胞 高表达 转染 骨发生 酶技术 

分 类 号：R346[医药卫生—基础医学]