Construction of mrp1 Expression Vector and Biological Characteristics in HepG2 Cells_CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Authors：

WANG Xinping ¹ , YAN Lnan ¹ , PAN Guangdong ¹ , XIA Dong ¹ , ZHANG Mingman ¹ , GOU Xinhua ² , ZHAO Lanying ²

Department of General Surgery, West China Hospital, Sichuan University, Chengdu 610041, China;

Corresponding author：

Keywords：

Multidrug resistance Multidrug resistance-associated protein Human hepatocellullar carcinoma Plasmid

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【Abstract】ObjectiveTo construct a mrp1 expression vector and investigate its biological characteristics in HepG2 cells in vitro.
MethodsThe 6.5 kb multidrug resistanceassociated protein （MRP) cDNA obtained from plasmid pGEM-mrp1 was cloned into the pCI-neo mammalian expression vector, which was later transferred into human hepatocarcinoma cell line HepG2 by liposome. Then the HepG2 cells resisting G418 were clustered and proliferated, and the mrp1 mRNA and MRP in these HepG2 cells were detected by means of RT-PCR and FCM respectively.
ResultsThe mrp1 expression vector was established successfully, and the stable MDR hepatocarcinoma cell line (HepG2/mrp1) was developed as well. The content of the specific fragment of mrp1 mRNA was (56.8±6.37)% and MRP was 7.89 in the HepG2/mrp1 cells, the corresponding value in HepG2 cells was (9.67±3.26)% and 0.79 respectively. The difference was statistically significant (P＜0.05).
ConclusionIt is practicable to establish MDR hepatocarcinoma cell line by transferring mrp1 cDNA into HepG2 cells, which is useful in the research of MDR mechanism.

Citation： WANG Xinping ,YAN Lnan,PAN Guangdong,XIA Dong,ZHANG Mingman,GOU Xinhua,ZHAO Lanying. Construction of mrp1 Expression Vector and Biological Characteristics in HepG2 Cells. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2006, 13(2): 163-166. doi: Copy

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2. Nagata J， Kijima H， Hatanaka H， et al. Reversal of drug resistance using hammerhead ribozymes against multidrug resistanceassociated protein and multidrug resistance 1 gene [Ｊ]. Int J Oncol， 2002; 21(5)∶ 1021.
3. Lage H, Perlitz C, Abele R, et al. Enhanced expression of human ABCtransporter tap is associated with cellular resistance to mitoxantrone [Ｊ]. FEBS Lett， 2001； 503(2-3)∶179.
4. Stavrovskaya AA. Cellular mechanisms of multidrug resistance of tumor cells [Ｊ]. Biochemistry， 2000； 65(1)∶95.
5. Ruefli AA, Smyth MJ, Johnstone RW. HMBA induces activation of a caspaseindependent cell death pathway to overcome Pglycoproteinmediated multidrug resistance [Ｊ]. Blood， 2000； 95(7)∶2378.
6. Loe DW, Deeley RG, Cole SPC. Biology of the multidrug resistanceassociated protein, MRP [Ｊ]. Euro J Cancer,1996； 32A（6）∶945.
7. 姚辉华, 刘忠，严律南，等. 多药耐药相关蛋白基因在原发性肝细胞癌组织中的表达及意义 [Ｊ]. 中国普外基础与临床杂志， 2003； 10（6）∶576.
8. 戴越盟，林萍. 多药耐药相关蛋白在阿霉素诱导人肝癌细胞SMMC7721耐药性产生中的作用及机理 [Ｊ]. 中国普外基础与临床杂志， 2001； 8（1）∶8.
9. Mavel S, Dikic B, Palakas S, et al. Synthesis and biological evaluation of a series of flavone derivatives as potential radioligands for imaging the multidrug resistanceassociated protein 1 (ABCC1/MRP1) [Ｊ]. Bioorg Med Chem， 2005； 28(4)∶521.
10. Godinot N, Iversen PW, Tabas L, et al. Cloning and functional characterization of the multidrug resistanceassociated protein (MRP1/ABCC1) from the cynomolgus monkey.
11. 陈永兵，余少鸿，严律南，等. 多药耐药真核表达载体的构建及其在人肝癌细胞HepG2中表达 [Ｊ]. 中国普外基础与临床杂志， 2005； 12（3）∶254.
12. 翟宝进，伍烽，邵泽勇, 等. 阿霉素诱导人肝癌细胞多药耐药株的建立及其生物学特性评价 [Ｊ]. 癌症， 2004； 23（4）∶391.
13. 邵泽勇，沈鼎明，伍烽, 等. 人肝癌原发性耐药细胞亚系的建立及耐药机制的初步探讨 [Ｊ]. 肿瘤, 2004； 24（5）∶455.
14. 顾玲，刘霆，龚玉萍. Mdr1单因素耐药白血病细胞株的构建 [Ｊ]. 四川大学学报（医学版）， 2005； 36（4）∶475.
15. 章小平，杨红枚，鲁功成. 膀胱癌MRP亚克隆的建立及其MDR表型 [Ｊ]. 中华肿瘤杂志， 2000； 22（4）∶273.

CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Construction of mrp1 Expression Vector and Biological Characteristics in HepG2 Cells

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