Experiment of the effect and safety of mdr1 gene transferring into k562 cells

WANG Shan; et al

doi:10.13406/j.cnki.cyxb.2003.03.011

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Experiment of the effect and safety of mdr1 gene transferring into k562 cells

更新时间：2024-01-17

- Experiment of the effect and safety of mdr1 gene transferring into k562 cells
- Journal of Chongqing Medical University Issue 3, Pages: 292-295(2003)
- 作者机构：
  
  1. 重庆医科大学儿科医院外科
  2. 重庆医科大学儿科医院外科,重庆,400014
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.2003.03.011
  CLC： R73-36
- Published：2003，
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[1]王珊,金先庆,杨纯正.mdr1基因转染K562细胞株的高效及安全性实验[J].重庆医科大学学报,2003(03):292-295.

WANG Shan, et al. Experiment of the effect and safety of mdr1 gene transferring into k562 cells. [J]. Journal of Chongqing Medical University (3):292-295(2003)
[1]王珊,金先庆,杨纯正.mdr1基因转染K562细胞株的高效及安全性实验[J].重庆医科大学学报,2003(03):292-295. DOI： 10.13406/j.cnki.cyxb.2003.03.011.

WANG Shan, et al. Experiment of the effect and safety of mdr1 gene transferring into k562 cells. [J]. Journal of Chongqing Medical University (3):292-295(2003) DOI： 10.13406/j.cnki.cyxb.2003.03.011.

摘要

目的 :mdr1基因转染K5 6 2细胞株

了解转染的持续稳定性、功能性及安全性。方法 :磷酸钙沉淀法建立一株双向性产病毒包装细胞

浓缩病毒上清液法感染K5 6 2细胞

分别用集落筛选法、流式细胞仪及SP免疫组化法检测转染率

柔红霉素泵出实验及MTT法测定外源性Pgp功能

流式细胞仪测细胞周期及PI分析

免疫组化测bcl- 2、c -myc的表达

柱形图、折线图及配对t检验进行统计学分析。结果 :①最高转染率为 34%

秋水仙碱筛选后达 84 %。②外源性mdr1基因可持续表达4月多

但表达率及强度逐渐降低。③转染的mdrl基因产物Pgp具有外排泵功能。④被转染的K5 6 2细胞获得 1.4 6～ 2 .2 2倍的多药耐药功能。⑤转染行为对靶细胞生长、增殖、凋亡呈一过性、轻度的、可逆的影响。结论 :mdrl基因转染K5 6 2细胞获得外源性Pgp高效、持久、功能、安全地表达

这为进一步研究mdrl基因修饰的骨髓移植在化疗中保护骨髓免受化疗损害奠定了基础。

Abstract

Objective:To transfer full-length human mdrlcDNA into K562 cells and assess the feasibility and safety in vitro.Methods:PA317 cells were transducted via a virus vector

pHaMDR1/A

containing full-length human mdr1cDNA by calcium phosphate precipitation.The K562 cells were infected with the virus supernatant.The transfection rate was determined by using CFU selection

FACS and SP immunohistochemical methods respectively.The foreign Pgp function was tested by MTT assay and DNR exclusion with FACS analysis .The cellular cycle and the expression of bcl-2 and c-myc gene were detected by using CFU sclcction

FACS and SP immunohistochemical mcthods respcctively.The forcign Pgp function was tested by MTT assay and DNR exclusion with FACS analysis.The cellular cycle and the expression of bcl-2 and c-myc gene were detected by using PI via FCM and SP stain respectively.These data were analyzed with statistic graph and T-test for match pairs.Results:①The highest transfection rate of K562/MDR6-18 cells were 34%

even up to 84% after colchicine selection.②The exogenous Pgp expression of K562/MDR6-18 cells lasted for more than 4 months with a slow decrease after 10 generations.③The exogenous Pgp got an exclusive pump function.④K562/MDR6-18 cells appeared 1.46-2.22 times cross multidrug resistance phenotype.⑤K562/MDR6-18 cellular proliferation showed a mild

short

restored change.There was no abnormal apoptosis or proliferation.Conclusions:It implies a feasible

attractive and safe way that mdr1 gene transfers into target cells to get exogenous effective MDR.They also provide the basic protocol for using mdr1 gene modifying bone marrow transplantation against the myelosuppressive effects of anticancer drugs.

关键词

mdr1基因K562细胞株转染凋亡

Keywords

Mdr1 geneTransfectionK562apoptosis

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