Construction of eukaryotic expression vector co-expressing genes encoding <em>T-bet</em> and <em>c-FLIP</em> and its biological function

YI Yuan-xue; LI Bao-jin; LIU Xiao-ping; ZHANG Chao; ZHANG Wei; FENG Zi-yi; LENG Xi-sheng

您当前的位置：

首页 >

文章列表页 >

Construction of eukaryotic expression vector co-expressing genes encoding T-bet and c-FLIP and its biological function

更新时间：2025-01-15

- Construction of eukaryotic expression vector co-expressing genes encoding T-bet and c-FLIP and its biological function
- Journal of Army Medical University Vol. 30, Issue 24, Pages: 2272-2275(2008)
- 作者机构：
  
  1. 北京大学香港科技大学医学中心，北京大学深圳医院肝胆腔镜外科
  2. 北京大学人民医院肝胆外科
- 作者简介：
- 基金信息：
- DOI：
  CLC： R392.2;R392-33;R392.12
- Published：2008，
  
  Published Online：30 December 2008，
- 稿件说明：
移动端阅览
YI YUAN-XUE, LI BAO-JIN, LIU XIAO-PING, et al. Construction of eukaryotic expression vector co-expressing genes encoding T-bet and c-FLIP and its biological function. [J]. Journal of army medical university, 2008, 30(24): 2272-2275.
DOI：

YI YUAN-XUE, LI BAO-JIN, LIU XIAO-PING, et al. Construction of eukaryotic expression vector co-expressing genes encoding T-bet and c-FLIP and its biological function. [J]. Journal of army medical university, 2008, 30(24): 2272-2275. DOI：

摘要

目的构建携带

T-bet

和

c-FLIP

双基因的真核表达载体及检测其生物学活性。方法采用RT-PCR方法从人外周血淋巴细胞的总RNA中扩增出T-bet和c-FLIP cDNA，利用pIRES和pEGFP-C1，构建

T-bet

和

c-FLIP

双基因真核表达载体pEGFPT-bet-IRES-c-FLIP。采用非脂质体的脂质型介导载体将pEGFPT-bet-IRES-c-FLIP转染至外周血淋巴细胞，观察

T-bet

和

c-FLIP

在外周血淋巴细胞中的表达，并检测其诱导淋巴细胞的抗凋亡作用及Th1/Th2细胞偏移。结果成功构建pEGFPT-bet-IRES-c-FLIP质粒，运用非脂质体的脂质型介导载体将该质粒转染淋巴细胞，其转染率为34.6%。流式细胞仪分析未经pEGFPT-bet-IRES-c-FLIP预处理的淋巴细胞在CH-11作用24 h后，细胞的凋亡率分别为（50.12±8.02）%；经pEGFPT-bet-IRES-c-FLIP预处理1 d的淋巴细胞，其细胞凋亡率分别下降到（5.73±0.37）%；双基因表达的淋巴细胞的Th1型细胞因子IFN-γ表达水平明显高于空载体表达的淋巴细胞及未经处理的淋巴细胞组（

0.01），且Th2型细胞因子IL-4表达水平较空载体表达的淋巴细胞及未经处理的淋巴细胞低（

0.01），差异有统计学意义。结论转染

T-bet

和

c-FLIP

共表达基因后的T淋巴细胞可产生抗凋亡作用及向Th1细胞分化。

Abstract

Objective To construct a eukaryotic expression vector co-expressing genes encoding

T-bet

and

c-FLIP

and to detect its bioactivity in transfected PBMCs. Methods

c-FLIP

and

T-bet

genes were cloned from tota

l RNA of lymphocyte with RT-PCR methods

then constructed into the eukaryotic expression vector by using pIRES and pEGFP-C1 vectors. Lymphocyte from blood donated by hepatocellular carcinoma patients infected by pEGFPT-bet-IRES-c-FLIP with QuikFusin Transfection Kit induced the Th1/Th2 shift and produced anti-apoptosis derived from CH-11 antibody

whose anti-apoptosis evaluated with PI stain flow cytometry (FCM). Results

T-bet

and

c-FLIP

genes were successfully cloned from lymphocytes and incorporated into the eukaryotic expression vector. Exposed to CH-11 for 24 h

the apoptosis of lymphocyte preconditioned by pEGFPT-bet-IRES-c-FLIP for 24 h was significantly decreased

compared with no-preconditioned cells (5.73%±0.37%

50.12%±8.02%)

indicating that pEGFPT-bet-IRES-c-FLIP preconditioning protected lymphocyte against apoptosis induced by CH-11. Lymphocyte infected by pEGFPT-bet-IRES-c-FLIP significantly enhanced IFN-γ levels and decreased IL-4 levels

compared with no-preconditioned cells and the cells infected by empty vector. Conclusion A eukaryotic expression vector co-expressing genes encoding

T-bet

and

c-FLIP

may induce anti-apoptotic activities of lymphocyte and Th1 differentiation.

关键词

T-bet基因c-FLIP基因pEGFP-C1T淋巴细胞生物学活性共表达

Keywords

T-bet genec-FLIP genepEGFP-C1T lymphocytecoexpressionbioactivity

references

Views

下载量

CSCD

Alert me when the article has been cited

提交

Tools

Publicity Resources

Correlation between regulation of dendritic cells by nuclear transcription factor-κB and imbalance of T helper cell differentiation in pathogenesis of henoch-schonlein purpura during childhood

Correlation between expression of granzyme B and perforin and apoptosis in epithelial cells in oral lichen planus with local lymphocyte infiltration

Gene expression profiles of activated T lymphocyte with IBP deficient by oligonucleotide microarray

Clinical characteristics of coronavirus disease 2019 in a Chongqing hospital: analysis of 89 cases

Related Author

GAO Hongli

PANG Ke

HUANG Chengcheng

Yang Haiping

Li Cuicui

Wang Li

Zhang Gaofu

Wang Lijia

Related Institution

Department of Geriatrics, Ward of Geriatrics and Novel Coronavirus Pneumonia, Department of Ultrasonography, Yongchuan Hospital of Chongqing Medical University

Department of Kidney Immunity， Children’s Hospital, Chongqing Medical University

暨南大学：医学院口腔系，第一附属医院口腔内科，医学院病理学教研室

深圳市瑞尔齿科门诊部

Department of Clinical Biochemistry, College of Laboratory Medicine, Third Military Medical University

⁰

Construction of eukaryotic expression vector co-expressing genes encoding T-bet and c-FLIP and its biological function

DOI：

摘要

Abstract

关键词

Keywords

references