β-catenin依赖性LEF-1亚型对宫颈癌HeLa细胞生物行为学的影响

Effects of β-catenin-dependent LEF-1 isoforms on the biological behavior of HeLa cells

目的探讨β-catenin依赖性LEF-1亚型对宫颈癌HeLa细胞生物行为学的影响。方法利用PCR方法从人淋巴结cDNA文库中克隆全长型LEF-1的编码基因,插入到真核表达载体pcDNA3.1/V5-His中,脂质体法转染Hela细胞,G418筛选稳定表达目的基因的细胞株,Western blot鉴定目的基因的表达,MTT和平板克隆形成实验检测转染后细胞的增殖情况,Annexin V方法检测细胞凋亡情况,裸鼠体内成瘤实验检测β-catenin依赖性LEF-1亚型对肿瘤细胞体内成瘤能力的影响。结果成功构建LEF-1真核表达质粒,获得稳定表达β-catenin依赖性LEF-1亚型的HeLa细胞株,转染后的细胞增殖速度加快,凋亡水平降低,体内成瘤能力加强。结论全长型LEF-亚型的上调表达对于HeLa细胞的部分恶性生物学行为的发生具有一定的促进作用。

Objective To study the effects of β-catenin-dependent lymphoid enhancer factor（LEF-1） isoforms on biological behavior of HeLa cells.Methods β-catenin-dependent LEF-1 genes were obtained by PCR from human lymphoid node cDNA library and inserted into pcDNA3.1/V5-His vector to construct the eukaryotic expression plasmid pcDNA3.1-F-LEF-1.Using lipofectamineTM 2000,the plasmid pcDNA3.1-F-LEF-1 was transfected into Hela cells.Then we screened the stable cell lines that expressed the truncated LEF-1 isoforms by G418 and identified the expression of target gene with Western blot.Then we analyzed the proliferation,apoptosis,cell clone formation and capability of tumor formation in vivo of transfected cell lines.Results We successfully constructed the β-catenin-dependent LEF-1 eukaryotic expression plasmid and obtained the stable HeLa cell lines that expressed the full-length LEF-1 isoforms.The proliferation and capability of tumor formation in vivo of transfected cells were increased while apoptosis was decreased.Conclusion The overexpression of β-catenin-dependent isoforms can stimulate the malignant biological behavior of HeLa cells.