Mov10稳定表达肝癌细胞株的建立及其生物学研究  

Establishment of human hepatocellular carcinoma cell line stably over-expressing Mov10 and its biological behaviors

作　　者：刘敏慧[1] 童师雯[1] 盛云建[2] 殷文伟[1] 彭虹[1] 任红 汤慧[1,3] 

机构地区：[1]重庆医科大学:附属第二医院感染科,重庆加400010 [2]泸州医学院附属医院感染科,四川泸州646000 [3]病毒性肝炎研究所,重庆400010

出　　处：《第三军医大学学报》2014年第6期583-586,共4页Journal of Third Military Medical University

基　　金：国家自然科学基金重点项目(30930082);国家自然科学基金青年科学基金(81300258)~~

摘　　要：目的构建稳定表达Mov10蛋白的HepG2细胞株,探讨Mov10在肝癌细胞增殖和侵袭等生物学行为中的作用。方法将含Mov10基因的真核表达载体pCMV-Mov10-GFP质粒,采用脂质体介导法转染至人肝癌细胞系HepG2,经过G418抗性筛选阳性克隆。采用qPCR、Western blot技术检测Mov10的表达,免疫荧光技术观察稳定株Mov10的细胞分布。通过MTS、平板克隆形成试验、体外侵袭能力检测观察稳定细胞株的增殖情况、克隆形成能力和侵袭能力。结果成功构建了表达Mov10蛋白的稳定细胞株HepG2-Mov10。内源性及外源性Mov10蛋白主要分布于细胞质。稳定细胞株HepG2-Mov10较对照组细胞增殖速度快(P<0.05),体外侵袭能力强(P<0.05)。稳定细胞株HepG2-Mov10、HepG2-GFP和细胞株HepG2的克隆形成能力分别为(56.7±17.3)%、(38.3±12.1)%、(31.3±8.1)%,HepG2-Mov10细胞与后两者间差异有统计学意义(P<0.05)。结论成功构建稳定表达Mov10的细胞株HepG2-Mov10,发现Mov10能促进肝癌细胞增殖并提高克隆形成能力和侵袭能力。Objective To establish a cell line stably expressing Moloney leukemia virus 10 （Movl0） protein in HepG2 cells and investigate the role of Movl0 in the cell proliferation and invasion. Methods The eukaryotic expression plasmid pCMV-Movl0-GFP was transfeeted into the HepG2 cells mediated by lipo- fectamine and then selected with G418. The mRNA and protein expression of Movl0 was detected by qPCR and Western blotting, respectively. Immunofluorescence assay was used to analyze the location of Movl0. The proliferation and colony-formation ability was measured by MTS assay and plate colony-formation assay. Cell invasion assay was performed in vitro. Results Our established HepG2-Movl0 Cells stably expressed Movl0 protein. Endogenous and exogenous Movl0 protein was mostly distributed in the cytoplasm. HepG2-Movl0 ceils had significantly higher proliferative rate and stronger ability of invasion than HepG2 cells （P 〈 0. 05 ）. Colony forming efficiency was （56.7 ± 17.3 ） %, （38.3 ± 12.1 ） %, （31.3 ± 8.1 ） % respectively in HepG2-Movl0, HepG2-GFP and HepG2 cells, with the fromer significantly higher than the other 2 cell lines （P 〈 0. 05 ）. Conclusion A hepatoma stable cell line over-expressing Movl0 is successfully established. Stable overexpres-sion of Movl0 protein obviously promotes cell proliferation, colony formation and invasion.

关 键 词：癌 肝细胞 Mov10 质粒 稳定细胞株 

分 类 号：R73-351[医药卫生—肿瘤] R730.2[医药卫生—临床医学]