凋亡素基因对人结肠癌细胞作用的研究  

Experimental study on effect recombinant pEGP-VP3 on human colonic carcinoma cell line

作　　者：朱惠明[1] 王娜[1] 蔡筱彦[1] 邓传珍[1] 

机构地区：[1]暨南大学医学院第二附属医院深圳市人民医院消化内科,518020

出　　处：《现代消化及介入诊疗》2005年第3期125-128,共4页Modern Interventional Diagnosis and Treatment in Gastroenterology

摘　　要：目的探讨凋亡素基因的导入对人结肠癌细胞的影响。方法将凋亡素基因VP3克隆入真核表达载体pEGFP-C2,构建成重组质粒pEGFP-VP3。用脂质体转染法将pEGFP-C2和pEGFP-VP3分组分别转染人类结肠癌细胞(Lovo)和小鼠胚胎成纤维细胞(NIH3T3),通过荧光显微镜观察绿色荧光蛋白在不同组细胞中的定位、表达及细胞的生长、凋亡情况,MTT法测定不同组细胞的生长曲线,并用An-nexinV-FITC流式细胞技术检测细胞的凋亡率。结果在Lovo/EGFP组和3T3/EGFP-VP3组细胞中,EGFP均匀分布于细胞中,细胞形态无明显变化,细胞凋亡率较非转染细胞组亦无明显增加。而在Lovo/EGFP-VP3组细胞中,EGFP-VP3以荧光颗粒形式集中在细胞核,并逐渐变粗,最后细胞碎裂成片状,流式细胞技术检测Lovo/EGFP-VP3组的细胞凋亡率明显高于其他对照组(P<0.01)。结论pEGFP-VP3可诱导人类结肠癌细胞凋亡。Objective To investigate the effect of expression of apoptin gene on human colonic carcin- ma cells. Methods Recombinant plasmid pEGFP-VP3 was constructed by cloning an apoptin gene to eukaryotic expression vector pEGFP-C2. To investigate the effect of the apoptin （vp3） in human colonic carcinoma cells, two plasmids, pEGFP-C2 and pEGFP-VP3, were respectively transfected into Lovo （a carcinoma cell line came from human colon） and NIH3T3（a fibroblast cell line came from mouse embryo） cells, by a lipidsome lipid transfection method. The different green fluorescent cell in various groups, to infer the location and expression of EGFP/EGFP-VP3 and the growth or apoptosis state of cells, and the rate of apoptosis in various group cells was assayed by flow cytometer （FCM） with Annexin V-PI stain simultaneously. Results In the cells of Lovo/EGFP and 3T3/EGFP-VP3 group, EGFP was fairly well-distributed in whole cell. No change was found in cell morphology and apoptosis rate on transfected cells than natural ones. In the cells of Lovo/ EGFP-VP3 group, EGFP-VP3 was concentrated in nucleus as fluorescent granules, furthermore became coarser and the cells became small and round, and finally pieces. The rate of apoptosis in the cells of Lovo/ EGFP-VP3 group was obviously higher than other control group cells （P 〈 0.01）. Conclusion Gene VP3 is introduced into human colonic carcinoma cells which express protein apoptin and induce apoptosis.

关 键 词：凋亡素 增强型绿色荧光蛋白 人结肠癌 细胞凋亡 人结肠癌细胞 细胞作用 基因 小鼠胚胎成纤维细胞 脂质体转染法 绿色荧光蛋白 

分 类 号：R735.35[医药卫生—肿瘤] R454.1[医药卫生—临床医学]