| Summary: | 碩士 === 國防醫學院 === 微生物及免疫學研究所 === 93 === Japanese encephalitis (JE) is a mosquito borne viral disease of mankind. Currently, the licensed vaccines for JEV are inactivated virus vaccines derived from mouse brain or cell culture. It needs repeatedly parenteral boost to effectively induce the protective immunity and hence the hypersensitivity reaction may be occurred. Alternatively, oral vaccine using bacteria or liposome as a delivery system which having the advantages of self-administration at ease, low risk of contamination, and reduced cost, has been demonstrated to be a promising approach for the development of the new generation of JE vaccines. Recently antifungal antibiotic amphotericin B has gained much attention due to its ability in modulation of the immune responses. In our previous study, we have demonstrated that amphotericin B was capable to augment the immune response of oral Salmonella vaccine to the carried antigen. In this study, the effect of AmB on immune response to an oral liposomal JE vaccine was investigated. Firstly, a liposome composed of DSPC (distearoylphosphatidylcholine) and cholesterol at a molar ratio of 2:1 was made to entrap the recombinant NS1 (rNS1) protein from JEV, designated Lip-JNS1. The serological response and the protective efficacy of the resulting Lip-JNS1 vaccine were assessed in mouse model following oral administration. Our results showed that the rNS1 was expressed as an inclusion body in E. coli BL21/pET21b-JNS1, of which production yield was approximately 15.8 mg/l. The Lip-JNS1, with an average size of 0.5 μm by extrusion, had an estimate of encapsulation protein rate of 23 %. The stability study revealed that Lip-JNS1 was stable in mimic gastric fluid and intestinal medium and appeared to be stable at 4 ℃ than that at room temperature for storage. Mice orally given three dose of Lip-JNS1 vaccine (0.5 ml/mouse) at 14 days apart were capable of inducing detectable anti-NS1 sera IgG titers. Furthermore, the systemic immune response to JEV NS1 and the protective efficacy against JEV infection were remarkably enhanced by amphotericin B when it was orally administered before giving the Lip-JNS1 oral vaccine.
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