AS-mCLB1重组质粒体外抗肿瘤及化疗增敏作用  

Recombined Plasmid of Full-length Cyclin B1 Antisense cDNA Inhibits Lewis Lung Carcinoma Cell Proliferation and Renders Cells More Sensitive to Gemcitabine in Vitro

作　　者：张伶[1] 魏于全[2] 宋英[1] 蒋磊[1] 陈平[2] 陈县城[2] 李继承[1] 

机构地区：[1]浙江大学细胞生物学研究所,杭州310058 [2]四川大学华西医院肿瘤生物治疗国家重点实验室,成都610041

出　　处：《细胞生物学杂志》2007年第4期539-544,共6页Chinese Journal of Cell Biology

摘　　要：研究反义全长小鼠细胞周期蛋白B1重组质粒(pAS-mCLB1)体外抗肿瘤及化疗增敏作用。扩增后少量抽提并纯化pAS-mCLB1,通过脂质体将其转染入小鼠露易丝肺癌(LL/2)细胞,RT-PCR和Western印迹测定细胞内细胞周期蛋白B1的表达,观察转染后细胞形态变化,MTT法检测细胞增殖活性,流式细胞仪检测细胞周期及凋亡。细胞转染48h后,用化疗药物健择(gemcitabine;0.2μmol/L)处理24h,MTT法测定健择对细胞的杀伤作用。研究提示pAS-mCLB1转染后LL/2细胞形态明显异常,细胞内细胞周期蛋白B1表达显著下调,细胞周期阻滞于G1期,增殖受抑,凋亡增加;健择对pAS-mCLB1转染后LL/2细胞的杀伤作用显著增强。重组质粒pAS-mCLB1体外具有明显的抗肿瘤作用,并能增强肿瘤细胞对化疗药物的敏感性,估计上述作用与其下调肿瘤细胞内细胞周期蛋白B1表达,从而诱导细胞周期阻滞及凋亡等作用相关。Study the effect of recombined plasmid of full-length mouse cyclin B1 antisense cDNA （pASmCLB1） inhibitting tumor cell proliferation and rendering cells more sensitive to gemcitabine in vitro. After being amplified and purified, pAS-mCLB1 was transfected into Lewis lung carcinoma （LL/2） cells using lipofectamine 2000. mRNA content of mCLB1 was semi-quantitatively assayed using RT-PCR. Morphological changes of cells were viewed through inverted phase contrast and scanning electron microscope. The inhibition of cell proliferation was estimated by MTT assay. Cell cycle and apoptosis of LL/2 cells were determined through flow cytometry. After 48 h being transfected with pAS-mCLB1, cells were treated with gemcitabine （0.2 μnol/L） for 24 h, and then the cytotoxicity of gemcitabine was measured by MTT assay. Prominent G1 arrest and apoptosis were shown and abnormal morphology with inhibition of cell growth appeared in the cells transfected with pAS-mCLB1 in which evident down-regulation of mCLB1 was induced. Additionally, cytotoxicity of gemcitabine to LL/2 cells transfected with pAS-mCLB1 was more obvious than controls, pAS-mCLB1 could suppress tumor cell proliferation and increase cell sensitivity to gemcitabine in vitro, which may be associated with its ability to down-regulate the expression of mCLB1 and then induce G1 arrest and cell apoptosis in tumor cells.

关 键 词：反义全长cDNA 细胞周期蛋白B1 细胞周期阻滞 凋亡 化疗敏感性 

分 类 号：R730.5[医药卫生—肿瘤]