TFPI-2对人肝癌细胞生长增殖、凋亡及AFP合成的影响  

Effects of TFPI-2 on Proliferation,Apoptosis and AFP Expression of Hepatocarcinoma Cells

作　　者：秦晓林[1] 徐勇 范晓卿[1] 李武县[1] 匡文斌[1] 成凤[1] 涂植光[1] 

机构地区：[1]重庆医科大学检验医学院临床检验诊断学教育部重点实验室,重庆400016 [2]广东省深圳市坪山区人民医院,深圳518118

出　　处：《中国生物工程杂志》2011年第12期33-38,共6页China Biotechnology

基　　金：深圳市科技计划资助项目(200902013)

摘　　要：目的:探讨TFPI-2基因对人肝癌细胞Hep3B生长增殖、凋亡及甲胎蛋白AFP表达的影响。方法:将重组质粒PCDNA3.1-TFPI-2转染Hep3B细胞并经G418稳定筛选后,RT-PCR和Westernblot检测转染前后TFPI-2 mRNA和蛋白表达水平,采用CCK-8法、生长曲线观察TFPI-2对人肝癌细胞Hep3B生长增殖的影响,通过平板克隆形成实验观察单个细胞的增殖能力,RT-PCR检测AFP mRNA的表达,并用电化学发光法测定培养上清液中甲胎蛋白AFP含量,流式细胞仪检测细胞早晚期凋亡情况。结果:转染成功的Hep3B细胞检测到TFPI-2 mRNA和蛋白的表达;与转染空载体及未转染的细胞相比,转染TFPI-2的细胞生长增殖能力明显减弱;AFP mRNA表达抑制率为16.51%,AFP蛋白分泌明显低于对照组(P<0.01);流式细胞术检测转染TFPI-2的Hep3B细胞早期凋亡率明显增加(24.03%±7.28%vs 8.77%±3.66%)。结论:TFPI-2表达可显著抑制肝癌细胞生长和AFP的表达,同时还能诱导细胞早期凋亡。为进一步探讨靶向TFPI-2的肝癌基因治疗提供了实验依据。Objective： To investigate the effects of tissue factor pathway inhibitor-2 （TFPI-2） gene on the proliferation, apoptosis and expression of alpha fetoprotein of hepatocarcinoma cell line Hep3B. Methods： The recombinant vector of PCDNA3.1-TFPI-2 was transfected into Hep3B cells with liposome, and the cells were selected by G418. The expressions of TFPI-2 mRNA and protein were detected by RT-PCR and Western blot respectively. The influence of TFPI-2 on the proliferation of Hep3B cells via growth curve was assayed by CCK-8 method. The colony-forming unit assay was used to measure single cell self-replication ability. The expression of AFP mRNA was detected by RT-PCR and the AFP secretion of Hep3B cell was measured by electro- chemiluminescence （ECL） immunoassay. Furthermore, the apoptosis was tested by flow cytometry. Results： Expressions of mRNA and protein of TFPI-2 were identified in cells transfeeted by PCDNA3.1-TFPI-2. The growth rate and self-replication ability of Hep3B cells were significantly lower than those of the two control groups. The inhibition ratio of AFP mRNA expression was 16. 51%, and the AFP secretion of Hep3B cells transfected with PCDNA3.1-TFPI-2 was significantly lower than the control groups （P 〈 0. 01 ）. The rate of early apoptosis was significantly increased （24. 03% ±7.28% vs 8.77% ±3.66% ）. Conclusions： The expression of TFPI-2 can simultaneously inhibit the growth and expression of AFP of hepatocarcinoma cells as well as induce early apoptosis.

关 键 词：肝癌 组织因子途径抑制物-2 细胞增殖 甲胎蛋白 细胞凋亡 

分 类 号：Q819[生物学—生物工程]