禽流感H5N1灭活疫苗与不同纳米化佐剂联合免疫研究

Comparison of efficacy of nano-aluminum hydroxide and nano-MF59 adjuvanted inactivated avianinfluenza vaccines （ HSN1 ） in mice

目的观察禽流感H5N1灭活疫苗加不同佐剂以及纳米化佐剂免疫小鼠后产生的免疫应答的差异，同时观察免疫对异亚型病毒攻击后的保护情况。方法用H5N1灭活疫苗分别联合佐剂氢氧化铝、纳米化氢氧化铝、MF59和纳米化MF59通过腹腔注射方式免疫雌性BALB／c小鼠，同时分别以H5N1灭活疫苗免疫小鼠以及PBS腹腔注射处理小鼠作对照。采用ELISA方法分别对各组小鼠免疫后血清特异性IgG及其亚类IgGl、IgG2a水平进行检测，以PR8病毒鼻腔攻击后观察小鼠体重变化情况和生存率。采用t检验作组间比较。结果与PBS处理组相比，无论以何种方式免疫H5N1疫苗，免疫后特异性IgG及其亚类水平均明显升高（t=7．4004，P〈0．01），以联合MF59后诱导的特异性抗体水平最高。其中疫苗单独免疫组和联合M59免疫组IgG2a水平升高明显，联合纳米化MF59免疫小鼠后IgG2a水平有所下降，IgG1水平有所升高；联合铝佐剂组以IgG1升高为主。攻毒后各组小鼠体重均出现下降，但疫苗单独免疫小鼠以及疫苗与佐剂联合免疫小鼠于攻毒后期体重恢复接近正常或者正常。PBS处理组小鼠攻毒后全部死亡，佐剂联合免疫组小鼠存活率100％，而疫苗单独免疫小鼠存活率为70％。结论H5NI疫苗无论是单独免疫或是联合不同佐剂免疫均可诱导较高水平的特异性抗体产生，有非常好的免疫原性。HSN1灭活疫苗诱导的抗体亚类以IgG2a为主，MF59以及纳米化MF59也以诱导／gG2a亚类为主，但纳米化MF59可诱导更均衡的免疫应答。联合铝佐剂或纳米化铝佐剂免疫小鼠后均可诱导以IgG1亚类为主的抗体应答。联合两种佐剂均可对小鼠产生很好的异亚型保护。

Objective To observe the differences between nano-aluminum hydroxide and nano-MF59 adjuvanted inactivated avian influenza HSN1 vaccine, and to compare the protection of mice against PR8 virus challenge. Methods The BALB/c mice were intraperitonealiy injected with inactivated HSN1 vaccine with nano-aluminum hydroxide, nano-MF59 adjuvant, aluminum hydroxide or MF59 adjuvant. The mice injected with PBS or inactivated HSN1 vaccine only were used as control. The levels of specific IgG to H5N1 and its subclasses IgG1 and IgG2a in serum obtained from mice in each group after vaccination were assayed by ELISA kits. Weight loss and the survival rates were observed in mice after PR8 virus challenge. The t test was adopted in the comparison between vaccination groups and control group. Results As compared with PBS group, the level of IgG, IgG1 and IgG2a increased in mice in each vaccination group （t =7. 4004,P 〈0. 01 ）, particularly in the MF59 adjuvanted vaccine group, showing the highest level of specific IgG. The IgG2a level increased significantly in vaccine group and MF59 adjuvanted vaccine group. The IgG2a level decreased and IgG1 level increased in nano-MF59 adjuvanted vaccine group. The IgG1 level increased mainly in aluminum hydroxide adjuvanted vaccine group. After PR8 virus challenge, weight loss was found in mice in all groups, bat the weight of mice in all vaccination groups returned to near normal or normal later, and the mice in PBS group all died. The survival rates were 100% in all adjuvanted vaccine groups, and 70% in non-adjuvanted vaccine group. Conclusiolls The non-adjuvanted and adjuvanted vaccines have strong immunogenicity, and induce higher level of specific antibody. The H5N1 vaccine, MF59 and nano-MF59 induce mainly IgG2a antibody, but the nano-MF59 adjuvant can induce the balanced immune response type. The aluminum hydroxide and nano-aluminum hydroxide can induce mainly IgG1 antibody response in mice. Both the aluminum hydroxide and MF59 adjuvant can protect the mice against heterosubtypic influenza virus challenge.