质粒表达型shRNA对CHL-3A4转基因细胞系细胞色素P450 3A4基因表达的抑制作用  

Inhibitory effect of CYP450 3A4 gene expression and function by vector expressed small hairpin interfering RNA in CHL-3A4 cells lines

作　　者：陈杰[1] 赵立子[1] 王雪丁[1] 关溯[1] 邓颖[1] 袁芳[1] 黄民[1] 

机构地区：[1]中山大学药学院临床药理研究所,广东广州510080

出　　处：《中国临床药理学杂志》2006年第4期262-266,共5页The Chinese Journal of Clinical Pharmacology

基　　金：广州市重大项目-创新药物药理毒理评价研究技术平台资助项目(2003Z2-E4071)

摘　　要：目的研究质粒表达型shRNA对CHL-3A4转基因细胞系细胞色素P4503A4基因表达的抑制作用。方法设计并构建3条shRNA真核表达载体(CYP3A4Ⅰ、CYP3A4Ⅱ、CYP3A4Ⅲ),转染CHL-3A4转基因细胞,以空载体组及无关序列的shRNA表达载体组为阴性对照;转染后48h,Westernblotting观察RNA干扰对CYP3A4蛋白表达的影响;RNA干扰后,用HPLC法测定CHL-3A4转基因细胞的硝苯地平氧化酶的活性。结果CYP3A4Ⅲ表达载体能显著抑制CYP3A4蛋白表达(75%),抑制硝苯地平氧化酶的活性。结论RNA干扰能抑制CYP3A4基因表达。Objective To investigate the intlibitory effect of CYP 3A4 gene expression and function in CHL -3A4 cells lines by vector expressed small hairpin interfering RNA （shRNA）. Methods The shRNA expression vectors targeting CYP 3A4 gene （CYP3A4 Ⅰ , CYP3A4Ⅱ, CYP3A4 Ⅲ） were designed and constructed. The cells were transfected with shRNA expression vectors transiently, and the cells without shRNA transfection and with nonspecific shRNA transfection were used as controls. The inhibitory effect of RNA interfering was detected by Western blot analysis. The activity of nifedipine oxydase in CHL - 3A4 was mea - sured by HPLC assay. Results CYP 3A4Ⅲ shRNA expressing vector significantly reduced the protein expression levels （75%） of the CYP 3A4 gene and significantly inhibited the activity of nifedipine oxydase in CHL - 3A4 cells. Conclusion Vector - based RNA could suppress CYP 3A4 gene expression and function, and the use of RNA interfering interfering to inhibit CYP3A4 gene expression in mammalian cells was a promising new tool for the study of gene function.

关 键 词：细胞色素P450 3A4 代谢 硝苯地平 

分 类 号：R969.1[医药卫生—药理学] R972.3[医药卫生—药学]