國家衛生研究院 NHRI:Item 3990099045/1327
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    题名: Immunological characterizations of the nucleocapsid protein based SARS vaccine candidates
    作者: Liu, SJ;Leng, CH;Lien, SP;Chi, HY;Huang, CY;Lin, CL;Lian, WC;Chen, CJ;Hsieh, SL;Chong, P
    贡献者: Vaccine Research and Development Center
    摘要: The recombinant nucleocapsid (rN) protein of the coronavirus (CoV) responsible for severe acute respiratory syndrome (SARS) was cloned and expressed in Escherichia coli, extracted from cell lysates containing 6 M urea, then purified by Ni2+-affinity chromatography. In animal immunogenicity studies, we found that most anti-rN protein antibodies were IgG2a in BALB/c mice vaccinated with rN emulsified in Montanide ISA-51 containing the synthetic oligodeoxynucleotide, CpG. In contrast, anti-rN protein antibodies of mice immunized with rN protein in PBS were found to mainly be IgG1. These results indicated that ISA-51/CpG-formulated rN protein was dramatically biased toward a Th1 immune response. To identify the B-cell immunodominant epitopes of the rN protein in the mouse and monkey, the reactivities of antisera raised against purified rN proteins formulated in ISA-51/CpG were tested with a panel of overlapping synthetic peptides covering the entire N protein sequence. Three immunodominant linear B-cell epitope regions were mapped to residues 166-180, 356-375, and 396-410 of the rN protein. When the reactivities of these peptides were screened with human sera from five SARS patients, peptides corresponding to residues 156-175 reacted strongly with sera from two of the SARS patients. These results indicated that the region around residues 156-175 of the N protein is immunogenic in the mouse, monkey, and human. We found that peptides corresponding to residues 1-30, 86-100, 306-320, and 351-365 contained murine immunodominant T-cell epitopes. To identify functional CTL epitopes of the N protein, BALB/c mice were immunized with peptides containing the H-21K(d) CTL motif emulsified in adjuvant ISA-51/CpG. Using an IFN-gamma secretion cell assay and analysis by flow cytometry, peptides containing residues 81-95 were found to be capable of stimulating both CD4(+) and CD8(+) cell proliferation in vitro. We also only observed that peptides corresponding to residues 336-350 were capable of stimulating IFN-gamma production in T-cell cultures derived from peripheral blood mononuclear cells (PBMCs) of rnacaques immunized with the rN protein emulsified in ISA/CpG adjuvant. Our current results together with those of others suggest that some immunodominant B-cell and T-cell epitopes are conserved in the mouse, monkey, and human. This information is very important for the development SARS diagnostic kits and a vaccine. (c) 2006 Elsevier Ltd. All rights reserved.
    关键词: Immunology;Medicine, Research & Experimental;Veterinary Sciences
    日期: 2006-04-12
    關聯: Vaccine. 2006 Apr;24(16):3100-3108.
    Link to: http://dx.doi.org/10.1016/j.vaccine.2006.01.058
    JIF/Ranking 2023: http://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcAuth=NHRI&SrcApp=NHRI_IR&KeyISSN=0264-410X&DestApp=IC2JCR
    Cited Times(WOS): https://www.webofscience.com/wos/woscc/full-record/WOS:000236931900011
    Cited Times(Scopus): http://www.scopus.com/inward/record.url?partnerID=HzOxMe3b&scp=33645075167
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