Lentivector-mediated RNAi Efficiently Downregulates Expression of Murine TNF-α Gene in vitro and in vivo  

作　　者：王雪贞 唐荣华 薛峥 降风 张旻 卜碧涛 

机构地区：[1]Department of Neurology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology [2]Department of Neurology, Inner Mongolia Hospital

出　　处：《Journal of Huazhong University of Science and Technology(Medical Sciences)》2009年第1期112-117,共6页华中科技大学学报（医学英德文版）

基　　金：supported by grants from National Natural Sciences Foundation of China (No. 30670735, 30670737 and 30700244).

摘　　要：In order to explore the role of TNF-α in Niemann-Pick type C （NPC） disease, lentiviral-delivered RNA interference （RNAi） was used to silence the expression of murine TNF-α gene in vitro and in npc mice. Interference efficiency of the lentivirus expressing TNF-α-siRNA, previously constructed with the concentration of 2 x 108 ifu/mL, was determined by RT-PCR and ELISA in BV-2 cells and astrocytes. At the same time, the constructed Lenti-TNF-α-siRNA was intracerebroventricularly infused into 4-week old npc mice for a 4-week period, and the mice were divided into 3 groups： Lenti-TNF-α-siRNA （n=6）, control lentivirus （n=6）, and NPC mice without any intervention （n=4）. By using immunohistochemistry and real-time PCR, the down-regulation of the target genes was detected. The Lenti-TNF-α-siRNA downregulated the expression of murine TNF-α gene efficiently in vitro and the interference efficiency was 66.7%. Lentivirus could be expressed stably for long-term in the npc mice brain. Immunohistochemistry and real-time PCR revealed that, as compared with non-intervention group and Lenti-control group, Lenti-TNF-α-siRNA efficiently down-regulated the expression of murine TNF-α gene with the interference efficiency being 66.9%. TNF-α-siRNA downregulated the expression of TNF-α gene in vitro and in vivo, which provided a potential tool for studying and treating neurodegenerative diseases and TNF-α-related diseases.In order to explore the role of TNF-α in Niemann-Pick type C （NPC） disease, lentiviral-delivered RNA interference （RNAi） was used to silence the expression of murine TNF-α gene in vitro and in npc mice. Interference efficiency of the lentivirus expressing TNF-α-siRNA, previously constructed with the concentration of 2 x 108 ifu/mL, was determined by RT-PCR and ELISA in BV-2 cells and astrocytes. At the same time, the constructed Lenti-TNF-α-siRNA was intracerebroventricularly infused into 4-week old npc mice for a 4-week period, and the mice were divided into 3 groups： Lenti-TNF-α-siRNA （n=6）, control lentivirus （n=6）, and NPC mice without any intervention （n=4）. By using immunohistochemistry and real-time PCR, the down-regulation of the target genes was detected. The Lenti-TNF-α-siRNA downregulated the expression of murine TNF-α gene efficiently in vitro and the interference efficiency was 66.7%. Lentivirus could be expressed stably for long-term in the npc mice brain. Immunohistochemistry and real-time PCR revealed that, as compared with non-intervention group and Lenti-control group, Lenti-TNF-α-siRNA efficiently down-regulated the expression of murine TNF-α gene with the interference efficiency being 66.9%. TNF-α-siRNA downregulated the expression of TNF-α gene in vitro and in vivo, which provided a potential tool for studying and treating neurodegenerative diseases and TNF-α-related diseases.

关 键 词：LENTIVIRUS TNF-α neurodegenerative disease RNA interference npc mice 

分 类 号：R346[医药卫生—基础医学]