RNA干扰抑制马桑内酯诱导的大鼠星形胶质细胞Mdr1基因的表达  

Suppression of Mdr1 gene using RNA interference in cultured rat astrocytes induced by coriaria lactone

作　　者：杨天华[1] 程新望[1] 田林郁[1] 耿嘉[1] 陈蕾[1] 周东[1] 

机构地区：[1]四川大学华西医院神经内科,成都610041

出　　处：《中华神经科杂志》2007年第3期165-168,共4页Chinese Journal of Neurology

基　　金：国家自然科学基金资助项目(0040205401094)

摘　　要：目的观察 RNA 干扰对培养的大鼠星形胶质细胞逆转 P-糖蛋白介导的多药耐药现象的影响。方法用短发夹 RNA 表达载体 p-SIREN shuttle 质粒转染马桑内酯诱导表达 P-糖蛋白的星形胶质细胞,荧光定量聚合酶链反应(PCR)测定 Mdrl mRNA 的表达量,免疫细胞化学方法检测 P-糖蛋白表达的变化,流式细胞术检测转染前后细胞罗丹明蓄积外排的变化。结果建立表达 P-糖蛋白的星形胶质细胞模型,有效地将 p-SIREN shuttle 质粒转染该细胞模型。转染后实验组细胞 MdrlmRNA 水平被抑制达67.70%,P-糖蛋白表达明显低于对照组(P<0.01),实验组 P-糖蛋白作用底物罗丹明的细胞外泵出率(23.08%)明显低于对照组(78.35%,P<0.01)。结论 RNA 干扰对马桑内酯诱导的星形胶质细胞 Mdrl 表达有明显抑制作用,并且在很大程度上逆转多药耐药蛋白的耐药作用。Objective To reverse P-glycoprotein-mediated multiple drug resistance using RNA interference （RNAi） in cultured rat astrocytes. Methods Astrocytes overexpressing P-glycoprotein induced by coriaria lactone were transfected with the short-hairpin RNA expression vector-p-SIREN shuttle designed to target Mdrl mRNA. The mRNA level of Mdrl gene was evaluated by real-time PCR; the P-glycoprotein was examined by immunocytochemistry and image analysis, meanwhile the rhodamine efflux was assessed by flow cytometry. Results The astrocyte model overexpressing P-glycoprotein were established and successfully transfected with the short-hairpin RNA expression vector-p-SIREN shuttle. The mRNA level of Mdrlgene was knocked down by 67.70% （P 〈0. 01 ）. The expression of P-glycoprotein in the experimental group was significantly lower than that in negative control, and the rhodamine efflux ratio of experimental group （23. 08% ） was remarkably lower than that of negative control （78. 35%, P 〈0. 01 ）. Conclusions RNAi can knock down the expression of Mdrl in cultured rat astrocytes induced by coriaria lactone and then significantly reverse drug resistance. This methodologic strategy may pave a new way of further research of refractory epilepsy.

关 键 词：P糖蛋白 癫痫 星形细胞 内酯类 RNA 小分子干扰 

分 类 号：R742.1[医药卫生—神经病学与精神病学]