摘要
目的:观察阳离子脂质体LipofectamineTM 2000介导的寡脱氧核糖核苷酸(ODNs)在体外培养的人视网膜色素上皮(RPE)细胞中的转染效率及其在细胞中的分布。方法:在阳离子脂质体LipofectamineTM 2000的介导下将人工合成的FAM荧光标记的ODNs转入人RPE细胞中,荧光显微镜下观察转染后20 min、1 h、2 h、4 h、6 h、12 h、24 h其在细胞内的分布,流式细胞仪检测其在RPE细胞中的转染效率;并用MTT法检测脂质体处理组、脂质体-ODNs复合物处理组和正常对照组的细胞增生活性,观察脂质体的毒性。结果:LipofectamineTM 2000能迅速将FAM-ODNs转入人RPE细胞的胞质和胞核内,4~6 h达高峰,转染效率高达(85.85±5.75)%,与其他时点比较差异有统计学意义(P<0.05),24 h后其转染率还有(70.11±5.81)%;且脂质体对RPE细胞无明显细胞毒性,3组间细胞增生活性比较,差异无统计学意义(P>0.05)。结论:LipofectamineTM 2000能有效地将FAM-ODNs转入人RPE细胞中,可作为一种安全有效的基因转移载体用于基因治疗。
Aim: To investigate the transfection efficiency and distribution of oligodeoxynuclotide in human retinal pigment epithelial cells mediated by cationic liposome LipofectamineTM 2000.Methods:Fluorescence FAM labeled oligodeoxynuclotide was introduced into RPE cells by LipofectamineTM 2000.The location of fluorescence in RPE cells at 20 min,1 h,2 h,4 h,6 h,12 h,and 24 h after transfection was observed by fluorescence microscope,transfection efficiency was detected by flow cytometry,and the cytotoxicity was measured by MTT assay.Results: FAM-ODNs complexed with LipofectamineTM 2000 were transfected into cytoplasm and nucleus of RPE cells rapidly,and the highest transfect rate reached(85.85±2.75)% after 4 to 6 h incubation(P〈0.05),(70.11±5.81)% after 24 h.No cytotoxicity was observed(P〉0.05).Conclusion: LipofectamineTM 2000 could effectively transfect FAM-ODNs into RPE cells and it is a safe and effective vector for gene therapy.
出处
《郑州大学学报(医学版)》
CAS
北大核心
2008年第6期1111-1114,共4页
Journal of Zhengzhou University(Medical Sciences)