Development of DNA Vaccine Against Japanese Encephalitis Virus and Studies of Protective Mechanism

• Main Authors: Chien-Hsiung Pan, 潘建雄
• Other Authors: Mi-Hua Tao
• Format: Others
• Language: zh-TW
• Published: 2001
• Online Access: http://ndltd.ncl.edu.tw/handle/27976437663209592710

博士 === 國防醫學院 === 生命科學研究所 === 89 === In this study, we evaluated the protective role of the structural and nonstructural proteins of the Japanese encephalitis virus (JEV) in mice model by DNA immunization and demonstrated a high level of protection against a lethal viral challenge was elicited by immunization with a plasmid (pE) encoding the JEV envelope protein, whereas all the other genes tested, including those encoding the capsid protein and the nonstructural proteins NS1-2A, NS3 and NS5, were ineffective. Plasmid pE administered either by intramuscular or gene gun injection produced significant E-specific antibodies, helper T (Th) cell proliferative responses and CTL activities. A strong correlation between anti-E antibodies and the protective capacity was observed and confirmed by transfer experiments and gene knockout mice. Transfer of E-specific antiserum but not immune splenocytes to sublethally irradiated hosts conferred protection. Furthermore, experiments on gene knockout mice showed that DNA vaccination did not induce anti-E titers and protective immunity in Igm-/- and I-Ab-/- mice, whereas in CD8a-/- mice the pE-induced antibody titers and protective rate were comparable to those produced in the wild-type mice. We also test the concept of combining JEV E and nonstructural protein genes and demonstrated that immunization with plasmids encoding E and each one of NS1-2A, NS3 or NS5 genes gave more strong protection than them did alone. A significant enhancement of the anti-E antibodies, CTL response and IFN-g production was observed in mice received the co-administration of plasmids encoding E and nonstructural proteins. Among these different combinations, co-delivery with pE and NS1-2A gene provided the most protective capacity, and passive transfer with sera but not spleen cells conferred the complete protection. In addition, the immunization with pE plus pNS3 also provided the protection against a more stringent intra-cerebral challenge. We also investigated the protective immunity following the co-delivery of pE with plasmid encoding either of Th1 cytokine (IL-2 and IL-12), GM-CSF or Th2 cytokine (IL-4) genes. We observed dramatic enhancement of antigen-specific humoral response with the co-delivery of IL-4 and GM-CSF and increase of anti-E IgG2a in response to the co-delivery of Th1 cytokines. Furthermore, we found the similar humoral response induced by pE immunization between wildtype and IFN-g-/- and IL-4-/- mice, but a reversed Th2 and Th1 response was observed in mice immunized by intramuscular or gene gun injection. In addition, the complete lose of protection induced by pE with gene gun injection in IFN-g-/- mice was suggested the IFN-g_ is involved in the protective mechanism against JEV infection. Taken together, these results demonstrate that immunization with plasmids encoding JEV E protein alone or plus other nonstructural proteins can provide comparable or better protection than currently used vaccine. The protective mechanism of this DNA vaccine is mediated by Th cell-dependent antibody response, and IFN-g is also involved in this mechanism. Our studies provide the important basis for the development of more rationally designed JEV vaccine.