Inhibitory effect of RNAi on Japanese encephalitis virus replication in vitro and in vivo.
Flaviviruses include many insect-mediated small viruses and still cause serious problems in the world. In humans, JEV can cause acute meningioencephalomyelitis, resulting in fatality rates of 5 to 40%. RNA-interference (RNAi) as an antiviral mechanism was originally discovered in plants and then found in the specific suppression of gene expression of other organisms such as Caenorhabditis elegans, Drosophila and vertebrates. As JEV is an RNA virus, RNAi could be a reasonable approach for therapeutic purposes to use against Japanese encephalitis. In this study, we examined the effect of RNAi on JEV replication. Viral reproduction in Vero cells was decreased to 7.2% and 39.0% of control by the transfection of small interference RNAs, JCR and JN3R at 250 n M, respectively. Under the transfection of 5 microg/ml pJRi which produces stem-loop RNAi, viral reproduction was decreased to about 10% of control. Western blot analysis indicated that RNAi inhibited the translation level. We used pJRi in the animal experiment. After the inoculation of viruses at 5 x 10(3) PFU, pJRi at 1.0 and 5.0 microg/g was injected into mice i.p. JEV-infected control mice (n=5) died within 15 days. pJRi (1.0 or 5.0 microg/g)-medicated mice survived 40 or 80% at 15 days. The data clearly indicate that pJRi has highly potent inhibitory activity against JEV replication in vivo. The results in vivo and in vitro provide evidence that JEV replication was efficiently inhibited by RNAi and RNAis could be used as an antiviral drug against JEV infection.
Murakami
Manabu
M
Division of Molecular Oncology and Virology, Medical Research Institute, Kanazawa Medical University, Ishikawa 920-0293, Japan.
Ota
Takahide
T
Nukuzuma
Soichi
S
Takegami
Tsutomu
T
D000818:Animals; D002522:Chlorocebus aethiops; D004664:Encephalitis Virus, Japanese / Q000502:physiology*; D004672:Encephalitis, Japanese / Q000628:therapy*; D005260:Female; D006801:Humans; D051379:Mice; D008813:Mice, Inbred ICR; D020365:RNA Helicases / Q000235:genetics; D034622:RNA Interference; D012697:Serine Endopeptidases / Q000235:genetics; D014162:Transfection; D014709:Vero Cells; D017361:Viral Nonstructural Proteins / Q000235:genetics; D015678:Viral Structural Proteins / Q000235:genetics; D014779:Virus Replication / Q000235:genetics; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D020365:RNA Helicases; D012697:Serine Endopeptidases; D017361:Viral Nonstructural Proteins; D015678:Viral Structural Proteins; C060028:NS3 protein, flavivirus; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases