阳离子脂质体DOTAP佐剂对H5N1流感病毒裂解疫苗免疫效果的影响  

Effect from cationic liposomes DOTAP as adjuvant on H5N1 influenza split vaccine

作　　者：赵巍[1] 喻刚[1] 郝鹏亮[1] 韩锡鑫[1] 黄晓媛[1] 杨晓明[1] 

机构地区：[1]武汉生物制品研究所有限责任公司病毒性疫苗研究二室,湖北武汉430207

出　　处：《微生物学免疫学进展》2016年第2期1-9,共9页Progress In Microbiology and Immunology

基　　金：国家重大新药创制项目(2014ZX09102045-011)

摘　　要：目的研究阳离子脂质体DOTAP佐剂对H5N1型流感病毒裂解疫苗免疫效果的影响。方法制备DOTAP阳离子脂质体流感病毒裂解疫苗（简称DOTAP流感裂解疫苗）,检测其包封率。将BALB/c小鼠分为13组,分别用含0.1、1.0、10.0μg HA/只剂量以DOTAP、Al（OH）3、CPG-ODN为佐剂以及不含佐剂的流感裂解疫苗于0、21天皮下免疫,PBS作为对照组,用血凝抑制试验检测小鼠初次免疫后21、42天血清HI抗体滴度;用ELISA检测初次免疫后21、42天血清特异性IgG抗体、IgG1、IgG2a亚类抗体滴度,以及初次免疫后42天小鼠脾脏单个核细胞体外经抗原刺激后细胞因子IL-2、IL-4、IFN-γ的分泌水平。将BALB/c小鼠分为3组,分别用含不同DOTAP剂量（100、300、600μg/只）的DOTAP流感裂解疫苗于0、21天皮下免疫,检测初次免疫后21、42天小鼠血清HI抗体滴度和IgG抗体滴度。结果 DOTAP流感裂解疫苗粒径在300-400 nm,带正电荷,包封率在50%以上;DOTAP流感裂解疫苗诱导的HI抗体水平和特异性IgG抗体水平均高于流感裂解疫苗,而与铝佐剂和Cp G-ODN佐剂间差异无统计学意义;DOTAP流感裂解疫苗产生的抗体仍以IgG1亚类抗体为主,免疫后42天诱导的IgG2a亚类抗体水平高于流感裂解疫苗和铝佐剂,低于Cp G-ODN佐剂;DOTAP流感裂解疫苗免疫后既分泌高水平Th1型细胞因子IFN-γ,同时也分泌高水平Th2型细胞因子IL-4;不同DOTAP剂量的DOTAP流感裂解疫苗免疫后,其HI抗体滴度和IgG抗体滴度在低、中、高剂量组之间存在明显的量效关系。结论 DOTAP作为H5N1型流感病毒裂解疫苗的佐剂可显著提高流感裂解疫苗的免疫原性,其对体液免疫应答的增强作用不低于铝佐剂和Cp G-ODN佐剂,并具有诱导细胞免疫应答的能力。Objective To validate the enhanced immune responses when cationic liposomes DOTAP was used as the adjuvant in H5N1 influenza split vaccine（ simplified as influenza split vaccine）. Methods DOTAP cationic liposomes influenza vaccines were prepared by evaporating and hydrating of lipid films. A total of 13 groups of BALB / c mice were inoculated subcutaneously with 0. 1 μg,1. 0 μg,10. 0 μg HA dose in influenza split vaccine containing DOTAP,Alum CPG-ODN as adjuvant,respictively,and influenza split vaccine alone as negative control or PBS as blank at 0,21 day. Hemagglutinin inhibition（ HI） antibody titers were measured at 21,42 day,the specific Ig G antibody and subclass antibody Ig G1 and Ig G2 a were measured by ELISA at 42 day; and the single spleen cell was isolated and stimulated with antigen in vitro,then the released level of IL-2, IL-4, IFN-γ were measured by ELISA at 42 day. A total of 3 groups of BALB / c mice were inoculated subcutaneously by influenza split vaccines wsith 100,300,600 μg dose of DOTAP liposomes at 0,21 day. HI antibody titers were measured at 21,42 day,the specific Ig G antibody were measured by ELISA. Results All cationic liposome preparations had a diameter in 300- 400 nm with a postive charge,and reached more than 50% efficiency of encapsulation. Serology test indicated that the titers of HI antibody and specific antibody of DOTAP cationic liposomes influenza split vaccines were significantly higher than that of the influenza split vaccine without adjuvant,and had no a statistical difference with influenza split vaccine containing Alum or CPGODN. The higher level of Ig G1 antibodies were induced by DOTAP cationic liposomes influenza split vaccines in comparison of that of Ig G2 a antibodies. However,the antibody Ig G2 a titers were higher than that of the influenza split vaccine containig Alum in 42 day after vaccination,and lower than that of influenza split vaccine containing CPG-ODN. The result of Multiplex cytokine analysis indicated that both of high level of Th1 cytok

关 键 词：2 3-二油氧基丙基三甲基氯化铵 阳离子脂质体 佐剂 血凝抑制抗体 亚型抗体 细胞因子 

分 类 号：R392[医药卫生—免疫学]