ANGPTL4基因及缺失突变体对肝癌细胞生长的影响  

Effects of ANGPTL4 gene and its deletion mutants on growth of hepatocellular carcinoma cells

作　　者：王春艳[1] 李锦军[2] 张锋锐[2] 葛超[2] 万大方[2] 顾健人[2] 

机构地区：[1]复旦大学上海医学院,上海200032 [2]上海市肿瘤研究所癌基因及相关基因国家重点实验室,上海200032

出　　处：《肿瘤》2006年第3期219-223,共5页Tumor

基　　金：国家973项目(编号:2002CB513104);国家十五科技攻关重大项目(编号:2002BA711A02-1);上海市科委重点项目(编号:044119642;05JC14028)

摘　　要：目的:探讨ANGPTL4基因及其缺失突变体对肝癌细胞SMMC-7721生长的影响。方法:构建ANGPTL4基因全长及其功能结构域缺失突变体的融合表达载体,即ANGPTL4-GFP-N1、ANGPTL4-del1-GFP-N1、ANGPTL4-del2-GFP-N1、ANGPTL4-del3-GFP-N1,脂质体法将EGFP-N1空载体、ANGPTL4基因及其缺失突变体分别转染肝癌细胞SMMC-7721,G418筛选,建立稳定细胞系,用MTT试验法、细胞集落形成试验检测细胞体外生长活性和增殖能力;流式细胞仪检测ANGPTL4基因对细胞凋亡的影响;裸鼠移植瘤试验检测转染细胞的体内成瘤特性。结果:ANGPTL4基因及其缺失突变体体外对肝癌细胞SMMC-7721的生长、集落形成具有明显的抑制作用(均为P<0.01);ANGPTL4全长基因及其缺失突变体ANGPTL4-del1-GFP-N1和ANGPTL4-del2-GFP-N1均对SMMC-7721细胞体内成瘤有明显抑制作用(均为P<0.01)。但转染ANGPTL4全长基因及其缺失突变体的SMMC-7721细胞的细胞凋亡发生率与转染空载体组相比没有明显差别(均为P>0.05)。结论:ANGPTL4基因及其缺失突变体对肝癌SMMC-7721细胞体内、外生长均具有明显抑制作用,但该抑制作用不是通过促进细胞凋亡实现的。Objective;To explore the effect of angiopoietin-like 4 （ANGPTL4） gene and its deletion mutants on growth of hepatocellular carcinoma SMMC-7721 cells. Methods： ANGPTL40RF cDNA and its deletion mutants in functional domains（fibronectin like domain,signal peptide, coiled coil domain） were cloned into EGFP-N1 vector and named as ANGPTL4-GFP-N1, ANGPTL4 dell GFPN1, ANGPTL4-del2-GFP-N1, and ANGPTL4-del3-GFP-N1, respectively. Hepatocellular carcinoma SMMC-7721 cells were transfected with EGFP-N1 empty vector as control and ANGPTL4-GFP-N1, ANGPTL4-dell GFP N1, ANGPTL4-del2-GFP-NI,and ANGPTL4-del3-GFP-N1 by lipofectAMINE mediation. The stable colonies were obtained by G418 screening. The growth of SMMC-7721 cells were studied by in vitro colony formation assay and MTT assay. The ectopic tumor formation was assessed in tumor-bearing nude mice. Apoptosis and cell cycle of SMMC-7721 cells were analyzed by FACS. Results：Compared with control,transfection of ANGPTL4 and its deletion mutants significantly inhibited the growth and colony formation of SMMC-7721 cells in vitro （P〈0. 01）. Transfection of ANGPTL4 and its deletion mutants ANGPTL4-dell- GFP-N1 and ANGPTL4-del2-GFP-N1 also markedly inhibited the ectopic tumor formation of SMMC 7721 cells in nude Balb/c mice （all P〈0.01）. However,no significant difference in cell cycle and apoptotic ratio was observed between SMMC-7721 cells transfected with ANGPTL4/deletion mutants and those transfected with empty vector （P 〉0. 05, P 〉 0. 05 ）. Conclusion： ANGPTL4 and its deletion mutants can inhibit growth of SMMC-7721 cells in vitro and in vivo. Non apoptosis-mediated mechanisms are thought to contribute to the inhibitory effects.

关 键 词：肝细胞癌 ANGPTL4 突变 SMMC-7721细胞 

分 类 号：R735.7[医药卫生—肿瘤]