Twist在肝细胞癌中的表达及基因干扰Twist对细胞增殖和凋亡的影响  

Expression of Twist in hepatocellular carcinoma and effect of silencing Twist gene on the prolifera- tion and apoptosis of hepatocellular carcinoma cells

作　　者：刘永刚 刘瑾 龚铖[3] 

机构地区：[1]武汉市第十四医院肿瘤科,430200 [2]武汉市江夏区疾病控制中心 [3]武汉大学中南医院普外科

出　　处：《中华实验外科杂志》2014年第7期1501-1503,共3页Chinese Journal of Experimental Surgery

摘　　要：目的 探讨Twist在肝细胞癌中的表达及其与缺氧诱导因子（HIF）-1α的关系,观察Twist和HIF-1α基因干扰对肝细胞癌HepG2细胞增殖和凋亡的影响。方法 免疫组织化学法检测106例肝细胞癌组织中转录因子Twist和HIF-1α的表达,利用LipofectamineTM2000转染Twist和HIF-1α小分子干扰RNA（siRNA）至HepG2细胞。实验分5组：Twist干扰组、HIF-1α干扰组、双重干扰组、阴性对照组（转染negative-siRNA）和空白对照组（转染脂质体）,流式细胞仪检测细胞凋亡,噻唑蓝（MTT）比色法检测细胞体外增殖活力,染色质免疫共沉淀法（ChIP）检测Twist和HIF-1α基因间的直接作用关系。结果 癌组织Twist的阳性表达率为66.98%,明显高于癌旁组织的17.92%（P〈0.05）,Twist和HIF-1α的表达呈正相关（P〈0.05）。与空白对照组（3.98%）和阴性对照组（4.26%）比较,Twist干扰组、HIF-1α干扰组和双重干扰组细胞凋亡率分别为41.62%、43.84%和45.12%,干扰组细胞凋亡率均显著增加（P〈0.01）,且干扰组之间差异无统计学意义（P〈0.05）。Twist干扰组、HIF-1α干扰组和双重干扰组细胞生长抑制率分别为30.2%、34.0%和35.8%,各干扰组间差异无统计学意义（P〈0.05）。ChIP法证实在HepG2中Twist与HIF-1α启动子有直接作用。结论 Twist与HIF-1α基因表达呈正相关,表达下调后能诱导肝细胞癌HepG2细胞凋亡,并抑制其生长,双重干扰无明显协同作用,并且两基因间具有直接作用。Objective To investigate the expression of Twist in hepatocellular carcinoma （HCC）, the relationship between Twist and hypoxia inducible factor （ HIF）-1α, and the effect of silencing Twist and HIF-la genes on the proliferation and apoptosis of HCC HepG2 cells by RNA interference. Methods The expression of Twist and HIF-1a was detected in 106 cases of HCC by immunohistochemical staining method. Twist and HIF-1α-small interfering RNA （siRNA） were transfected into HepG2 cells mediated by LipofectamineTM 2000. HepG2 ceils were divided into five groups： Twist interference group, HIF-1α interference group, double interference group, negative control group （transfected with negative-siRNA） and blank control group （transfected with Lipofectamine）. Cell apoptosis was examined by flow cytometry and ceil growth was measured by methyl-thiazol-tetrazolium （MTr） assay. The direct interactions between Twist and HIF-1α were uncovered by chromatin immunoprecipitation assay （CHIP）. Results Twist posi- tive expression rate was high （66.98%） in HCC tissue but low in the corresponding peritumoral tissue （17.92%, P 〈0. 05）. There was a positive relationship between the expressions of Twist and HIF-1α （P 〈 0.05 ）. The apoptosis rate in blank control group, negative control group, Twist interference group, HIF-1α interference group and double interference group was 3.98%, 4. 26% , 41.62% , 43.84% , and 45.12%, respectively. The apoptosis rate in interference groups was significantly higher than that in control groups （ P 〈 0. 01 ）, while there was no significant difference among three interference groups （ P 〉 0. 05 ）. The cell growth inhibitory rate in Twist interference group, HIF-1α interference group and double interference group was 30. 2%, 34. 0% and 35.8% , respectively （ P 〉 0. 05 ）. There were the direct in- teractions between Twist and HIF-1α promoter in HepG2 by CHIP. Conclusion There was a positive relationship between the expression of Twist and HIF-1α

关 键 词：癌 肝细胞 TWIST 缺氧诱导因子-1Α 脱噬作用 细胞增殖 

分 类 号：R735.7[医药卫生—肿瘤]