沙门氏菌鞭毛蛋白对氢氧化铝制剂疫苗特异性免疫应答的影响

Enhanced immune response induced by the addition of Salmonella flagellin to an aluminium hydroxide preparation

目的评估沙门氏菌鞭毛蛋白作为添加剂在氢氧化铝疫苗制剂中的作用。方法以卵清蛋白(OVA)为抗原,氢氧化铝(Alhydrogel)为佐剂,鼠伤寒沙门氏菌鞭毛蛋白(FliC)或寡聚脱氧核糖核苷酸CpG-ODN 1826为添加剂,制备OVA/Alhydrogel、OVA/Alhydrogel+FliC和OVA/Alhydrogel+CpG-ODN 1826疫苗制剂。BALB/c小鼠随机分成3组,每组6鼠,分别用以上疫苗制剂以肌肉注射的方式免疫2次,间隔28d。在初次免疫后第28d和42d采小鼠血,分离血清,用ELISA法测定血清中抗OVA IgG及亚型。在初次免疫后的第42d处死小鼠,取脾,制备脾细胞,给予抗原刺激后检测培养上清中γ干扰素(IFN-γ)和白介素4(IL-4)水平。结果初次免疫后第42d,OVA/Alhydrogel+FliC制剂组OVA/Alhydrogel+CpG-ODN 1826制剂组和OVA/Alhydrogel制剂组特异IgG滴度分别为(714.75±213.32)U-nit、(863.78±355.05)Unit和(267.45±8.49)Unit,差异有统计学意义(P<0.05);前两制剂组比较差异无统计学意义(P>0.05)。IgG亚型分析和脾细胞培养上清IFN-γ和IL-4水平测定表明,OVA/Alhydrogel制剂诱导Th2偏向的免疫应答,而在加入沙门氏菌鞭毛蛋白后免疫应答有向Th1/Th2混合型转变的趋势。结论沙门氏菌鞭毛蛋白能增强特异性免疫应答反应,有望成为氢氧化铝疫苗制剂的添加剂。

Objective To evaluate the effect of Salmonella flagellin as an additive in an aluminium hydroxide preparation.Methods Ovalbumin（OVA） served as an antigen,aluminium hydroxide（Alhydrogel） served as an adjuvant,and phase I flagellin of Salmonella enterica serovar Typhimurium（FliC） or CpG-ODN 1826 served as an additive.Three vaccine preparations were prepared,namely,OVA/Alhydrogel,OVA/Alhydrogel＋FliC,and OVA/Alhydrogel＋CpG-ODN 1826.BALB/c mice were randomly divided into three groups of six and intramuscularly immunized two times with a 28-day interval.Mouse immune sera were collected on days 28 and 42 after the first immunization.Specific anti-OVA IgG antibody and subclass titers of the immune sera were assayed and analyzed using ELISA.On day 42,splenocytes were isolated from some immunized mice to detect the levels of two cytokines,interferon-γ（IFN-γ） and interleukin-4（IL-4）,secreted in response to stimulation with OVA.Results On day 42,the specific anti-OVA antibody titers induced by the respective vaccine preparations,OVA/Alhydrogel＋FliC and OVA/Alhydrogel＋CpG-ODN 1826,were（714.75±213.32） units and（863.78±355.05） units.Both were significantly higher than the titer generated by OVA/Alhydrogel,which was（267.45±8.49） units（P〈0.05）.The difference between the OVA/Alhydrogel＋FliC and OVA/Alhydrogel＋CpG-ODN 1826 groups was not statistically significant（P〉0.05）.The vaccine preparation of OVA/Alhydrogel induced a Th2 response.In contrast,the co-delivery of OVA/Alhydrogel and FliC promoted a Th2 response.This response tended to develop into a mixed Th1/Th2 response according to the results of analysis of subclass antibodies and the levels of INF-γ and IL-4.Conclusion Salmonella flagellin is a promising additive that can be used in aluminium hydroxide preparations to enhance antigen-specific immune responses.