TGF-β1通过RUNX2调控人肝星状细胞系LX-2中骨桥蛋白表达  

TGF-β1 Regulates the Expression of Osteopontin by Transcription Factor RUNX2 in LX-2 Cells

作　　者：王睿[1] 萧笑[1] 王红红[1] 赵丽娜[1] 徐立[1] 刘志国[1] 

机构地区：[1]第四军医大学西京消化病医院,陕西西安710032

出　　处：《现代生物医学进展》2013年第35期6801-6805,共5页Progress in Modern Biomedicine

基　　金：国家自然科学基金面上项目(81070350)

摘　　要：目的:探讨人肝星状细胞系LX-2中促纤维化因子TGF-β1对骨桥蛋白(Osteopontin,OPN)的转录调控作用,研究其潜在关系及作用通路。方法:经重组因子TGF-β1刺激后,western blot检测人肝星状细胞系LX-2中OPN和RUNX2的表达水平。通过生物信息学分析软件预测RUNX2在OPN启动子序列上的结合位点。构建基于pGL3-Basic的人OPN启动子载体和相应截短体,与RUNX2共转染HEK293细胞,结合双荧光素酶报告基因实验分析RUNX2对OPN启动子的转录调控作用。结果:经TGF-β1刺激后,LX-2细胞系中OPN与RUNX2的蛋白表达水平均升高,提示二者均为TGF-β1下游效应分子。TESS生物信息学分析显示OPN启动子序列-78^-73存在RUNX2的结合位点ACCACA。通过双荧光素酶报告基因实验结果显示:RUNX2对OPN启动子具有正向调控作用,且通过截短删除之前预测的结合位点后,该调控作用消失。结论:TGF-β1可促进人肝星状细胞系中OPN的表达,该作用部分通过其下游转录因子RUNX2作用于OPN启动子区域的ACCACA序列而实现。Objective： To investigate the regulation effect ofprofibrogenic factor TGF-β1 on osteopontin （OPN）. Methods： After LX-2 cell line was stimulated by recombinant TGF-β1, the levels of OPN and RUNX2 were detected by western blot. The binding site of RUNX2 on OPN promoter sequence was predicted by bioinformatic software, pGL3-Basic vectors containing OPN promoter sequence and truncated sequence deleting the predicted binding site were constructed, pGL3-OPN and RUNX2 were cotransfected into HEK293 cells, and the transcriptional effect of RUNX2 on OPN promoter was detected by dual luciferase reporter assay. Results： After the stimulation of TGF-β1, the levels of RUNX2 and OPN increased simultaneously, which indicated both molecules were downstream effector of TGF-β1 in LX-2 cell. Bioinformatic analysis demonstrated that a binding sequence ACCACA of RUNX2 located within OPN promoter region at -78--73. Dual luciferase reporter assay indicated that RUNX2 could promote the promoter activity of OPN and the effect disappeared when the predicted binding site was deleted in the truncated promoter sequence. Conclusions： TGF-β1 could promote the expression of OPN partly by RUNX2 acting upon ACCACA site located in OPN promoter sequence.

关 键 词：RUNX2 转录调控 骨桥蛋白 TGF-Β1 人肝星状细胞 

分 类 号：R575.2[医药卫生—消化系统]