复合HA-2氨基多肽可增强chitosan-DNA疫苗的免疫保护效果  

Complexing with HA-2-derived peptide (endosome fusogenic peptide) elevates the immunoprotection by chitosan-DNA vaccine

作　　者：徐薇[1] 沈燕[1] 陈瑞珍[2] 王缨[1] 熊思东[1] 

机构地区：[1]复旦大学上海医学院免疫学系教育部分子医学重点实验室上海基因免疫与疫苗研究中心,上海200032 [2]复旦大学附属中山医院卫生部病毒性心脏病重点实验室

出　　处：《中国免疫学杂志》2005年第2期93-98,共6页Chinese Journal of Immunology

基　　金：国家杰出青年科学基金 (3 992 5 0 3 1);国家重点基础研究发展计划资助项目 (2 0 0 1CB5 10 0 0 5 ) ;国家自然科学基金青年项目 (3 0 40 0 3 96)

摘　　要：目的 :以具有促黏膜黏附吸收功能的脱乙酰多糖chitosan包裹质粒DNApcDNA3-VP1构建的新型chitosan-DNA疫苗 ,已证实可诱生CVB3特异性黏膜IgA和抗CVB3保护力 ,在此基础上 ,引入具有内体破坏功能的流感病毒HA 2氨基多肽(融内体多肽 ) ,构建chitosan-DNA HApLys16疫苗 ,以期增强chitosan-DNA疫苗的免疫保护效果。方法 :将融内体多肽与多聚赖氨酸顺序合成为“载体多肽”HA pLys16 ,将其与DNA、chitosan依次复合形成chitosan DNA HApLys16疫苗。以 5 0 μgDNA剂量的chitosan-DNA疫苗滴鼻免疫BALB c小鼠 4次 ,检测CVB3特异性体液和细胞免疫应答 ;隔 4周以致死性 5×LD50 CVB3攻击小鼠 ,观察攻击后存活情况。以免疫组化法检测了小鼠心肌内CVB3载量 ;并检测了黏膜IgA中和效价的改变。结果 :chitosan DNA-HApLys16疫苗滴鼻免疫不仅诱生了高水平的IgG ,而且诱生了高水平的黏膜IgA抗体。其诱生的IgG水平以及特异性CTL杀伤活性与chitosan DNA疫苗无统计差别 ,而其IgA水平显著高于chitosan DNA疫苗。CVB3攻击后 ,chitosan DNA HAp Lys16可保护 5 0 .0 %小鼠长期存活 ,高于chitosan DNA组 4 2. 9%的免疫保护率。病理学研究显示 :chitosan-DNA HApLys16免疫小鼠心肌炎症状况好于chitosan-DNA免疫小鼠。Objective:To improve CVB3-specific immunity and protective ability of chitosan-DNA vaccine which was prepared by complexing chitosan, a kind of mucus absorption enhancer, with pcDNA3-VP1 and has been proved able to induce CVB3-specific mucosal IgA and anti-CVB3 protection.Methods:A novel non-viral “carrier peptide” system was introduced to chitosan-DNA vaccine which consist of two parts, one is poly-Lysine sequence to associate plasmid with functional peptide, and the other part is an endosome fusogenic peptide derived from HA-2 of influenza virus. A novel “viral-like” vaccine chitosan-DNA-HApLys16 vaccine was prepared by conjugating DNA with HApLys16 and then with chitosan. BALB/c mice were immunized intranasally 4 times with 50 μg DNA each time, and challenged with 5×LD_(50)CVB3 four weeks after the last immunization.Results:Intranasal immunization with chitosan-DNA-HApLys16 generated high levels of both IgG and IgA, the former was as the same level as induced with chitosan-DNA, while the latter was higher than that of chitosan-DNA immunized mice. CVB3 specific cellular immune response in terms of CTL activity was successfully induced and was not stronger than that of chitosan-DNA immunization. Chitosan-DNA-HApLys16 vaccine was able to protect 50% mice compared to 42.9% survival percentage purchased by chitosan-DNA vaccine. CVB3 particles stained with rabbit anti-CVB3 polyclonal antibody were not observed in chitosan-DNA-HApLys16 immunized mice but did exist in control mice. It was found that chitosan-DNA-HAplys16 induced the highest levels of IgA and the neutralizing titers of mucosal IgA turned out to be 1∶128, 1∶64 and 1∶8 for chitosan-DNA-HApLys16, chitosan-DNA and pcDNA3 group separately.Conclusion:Intranasal delivery of chitosan-DNA-HApLys16 vaccine could successfully promote the immunoprotection from 42.9% to 50% and this may be the result from higher mucosal IgA levels and also elevated neutralizing titers of IgA as seen in chitosan-DNA-HApLys16 immunization.

关 键 词：CVB3 clritosan-DNA HApLysl6 滴鼻 免疫保护 中和效价 

分 类 号：R392.12[医药卫生—免疫学]