TFPI-2对人肝癌细胞生长增殖、凋亡及AFP合成的影响

中国生物工程杂志

2011, Vol. 31

Issue (12): 33-38

研究报告

TFPI-2对人肝癌细胞生长增殖、凋亡及AFP合成的影响

秦晓林¹, 徐勇², 范晓卿¹, 李武县¹, 匡文斌¹, 成凤¹, 涂植光¹

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

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

QIN Xiao-lin¹, XU Yong², FAN Xiao-qing¹, LI Wu-xian¹, KUANG Wen-bin¹, CHENG eng¹, TU Zhi-guang¹

1. Key Laboratory of Laboratory Medical Diagnostics of Education Ministry, Chongqing Medical University, Chongqing 400016, China;
2. Pingshan People's Hospital, Shenzhen 518118, China

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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Western blot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。

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

Abstract:

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 transfected 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.

Key words: Hepatocarcinoma TFPI-2 Proliferation AFP Apoptosis

收稿日期: 2011-08-09 出版日期: 2011-12-25

ZTFLH:

Q819

基金资助:

深圳市科技计划资助项目(200902013)

通讯作者: 涂植光,电子信箱:tuzhiguang@yahoo.com.cn E-mail: tuzhiguang@yahoo.com.cn

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引用本文:

秦晓林, 徐勇, 范晓卿, 李武县, 匡文斌, 成凤, 涂植光. TFPI-2对人肝癌细胞生长增殖、凋亡及AFP合成的影响[J]. 中国生物工程杂志, 2011, 31(12): 33-38.

QIN Xiao-lin, XU Yong, FAN Xiao-qing, LI Wu-xian, KUANG Wen-bin, CHENG eng, TU Zhi-guang. Effects of TFPI-2 on Proliferation, Apoptosis and AFP Expression of Hepatocarcinoma Cells. China Biotechnology, 2011, 31(12): 33-38.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/ 或 https://manu60.magtech.com.cn/biotech/CN/Y2011/V31/I12/33

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