Lentivector-mediated RNAi efficiently downregulates expression of murine TNF-α gene in vitro and in vivo

Abstract

In order to explore the role of TNF-alpha in Niemann-Pick type C (NPC) disease, lentiviral-delivered RNA interference (RNAi) was used to silence the expression of murine TNF-alpha gene in vitro and in npc mice. Interference efficiency of the lentivirus expressing TNF-alpha-siRNA, previously constructed with the concentration of 2 x 10(8) ifu/mL, was determined by RT-PCR and ELISA in BV-2 cells and astrocytes. At the same time, the constructed Lenti-TNF-alpha-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-alpha-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-alpha-siRNA downregulated the expression of murine TNF-alpha 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-alpha-siRNA efficiently down-regulated the expression of murine TNF-alpha gene with the interference efficiency being 66.9%. TNF-alpha-siRNA down-regulated the expression of TNF-alpha gene in vitro and in vivo, which provided a potential tool for studying and treating neurodegenerative diseases and TNF-alpha-related diseases.