KAI1基因转染人乳腺癌细胞系的建立及初步研究  

Construction and Primary Study of Human Breast Cancer Cell Line Transfected by KAI1 Gene

作　　者：路丹[1] 王文秀[1] 徐玉清[1] 杨宇[1] 

机构地区：[1]哈尔滨医科大学附属第二临床医院肿瘤内科,黑龙江哈尔滨150086

出　　处：《现代生物医学进展》2007年第3期335-337,343,共4页Progress in Modern Biomedicine

基　　金：黑龙江省教育厅科学技术研究项目(No.11511235)

摘　　要：目的:通过将外源性KAI1基因转染入高转移性人乳腺癌细胞株,为乳腺癌基因治疗的实验室研究提供靶细胞,并初步探讨该基因对乳腺癌细胞增殖能力及细胞周期的影响。方法:采用脂质体法将pCMV-KAI1质粒转染入低表达KAIl基因的人乳腺癌细胞株MDA-MB-231中,经G418筛选后获得抗性克隆,利用RT-PCR、Western blot分析目的基因及其蛋白的表达情况,并利用MTT法和平板克隆形成实验初步探讨该基因对乳腺癌细胞体外增殖能力的影响,流式细胞术检测细胞生长周期的变化。结果:稳定转染KAI1基因的细胞株中有外源性目的基因和相应蛋白的高表达;MTT法示细胞增殖力下降,转染KAI1基因的集落形成率(25.33+2.36)％较转染前(43．17+2．75)％明显降低(P<0．05),流式细胞术显示转染KAI1基因后G1／G0期细胞数量由未转染前的(36．78+0．61)％升高至(64+7．56)％,M／G2期细胞数量则由(17．88+0．76)％降至(7．63+0．60)％,差异有显著性。结论:通过脂质体转染法获得了高表达KAI1基因及其蛋白的人乳腺癌细胞株,并发现该细胞株的体外增殖能力明显下降,这可能是KAI1基因通过调节细胞周期来实现的。Objective： To transfect the exogenous KAI1 gene to human breast cancer cell lines in order to provide the target cell in the study of gene therapy in breast cancer, and the influence of KAI1 gene on proliferative ability and the cycle pattern were studied. Methods： Human breast cancer cell line MDA-MB-231 was transfected with pCMV-KAI1 by lipofectamine and then selected by G418 to obtain the drug resistant monodone. RT-PCR and Western blot were used to determine the expression of KAI1 gene and its protein. The proliferative ability of cells was tested by MTF assay and colony-forming test and the cycle pattern were assayed by flow cytometry. Results： Cell clones ,stably transfected by gene KAI1, demonstrated the presence of exogenous target gene and over-expression of corresponding protein and their proliferative ability reduction was demostrated by MTF assay, also there was a significant reduction in colony-forming rate of pCMV-KAI1 transfected MDA-MB-231 cells （ （25.33 ＋2.36 ）% ） in comparison with the non-transfected ones （ （43.17 ＋2.75 ）% ） （P〈0.05）. Flow cytometry analysis revealed that in KAI1-transfected cell group the number of cell in G1/G0 phase increased to （64＋7.56）%, significantly higher than that of non-transfection （36.78＋0.61）%, while the number of cells in M/G2 phase significandy decreased from（17.88＋0.76）% to （7.63＋ 0.60）%. Conclusions： KAI1 gene can be expressed effectively in human breast cancer cell lines MDA-MB-231 after transfecfing the plasmid pCMV-KAI1 into the cell line by lipofectamine, with significant proliferative ability reduction of the transfected cell line, which is probably related to the change of the cell cycle pattern.

关 键 词：乳腺癌 KAIL基因 基因转染 增殖 

分 类 号：R737.9[医药卫生—肿瘤]