肿瘤相关新基因HC90的表达差异及初步功能研究

Differential expression and preliminary function study of novel tumor related cDNA clone HC90

目的 对新基因克隆HC90进行蛋白表达、各种肿瘤组织和细胞系中表达差异检测和体内外功能研究。方法 用pET 32a融合表达载体在大肠杆菌BL2 1(DE3)中对该基因进行表达 ,免疫小鼠制备小鼠源性多克隆抗体 ;从体外细胞集落形成试验 ,裸鼠体内成瘤实验 ,原位杂交 ,免疫细胞化学及免疫组化检测HC90基因的表达差异和对体内外细胞生长影响的生物学功能进行研究。结果 SMMC 772 1细胞系的体外集落形成试验表明 ,HC90转染组与空载体组和p5 3对照组相比集落形成数显著增加 (P <0 0 5和P <0 0 1) ;转染细胞系的裸鼠体内成瘤实验结果表明该基因对L92 9细胞系的成瘤有明显促进作用 (P <0 0 5 ) ;免疫组化结果显示HC90在人多种肿瘤组织 ,癌旁和正常组织中均有不同程度的表达 ,总体上表现为癌组织 >癌旁组织 >正常组织。结论 HC90是个新的肿瘤相关基因。

Objective The protein of novel cDNA clone HC90 was expressed. Differential expression in various tumors and cell lines and biological function of HC90 were studied in vitro and in vivo.Methods HC90 was expressed with fusion expression vector pET 32a in E.Coli BL21(DE3), and mouse anti HC90 fusion protein polyclonal antibody was prepared. The biologcial function and differential expression of HC90 were studied by in vitro colony formation assay, nude mouse tumor formation assay of transfected SMMC 7721 and L929 cell lines, in situ hybridization, immunocytochemistry and immunohistochemistry. Results Compared to pCMV Script vector and p53,HC90 accelerated the clony formation of transfected SMMC 7721 cell lines in vitro ( P <0 05 & P <0 01);Tumor formation assay of trasfected L929 cell line indicated that tumor formation was stimulated significantly ( P <0 05);immunohistochemistry showed that HC90 expressed in various tumor,noncancerous tissues and normal tissues to different extent. In general, HC90 expressed in human tissue as following extent:tumors>noncancerous tissues≥normal tissue.Conclusion HC90 is a novel tumor related gene.$$$$