鼠伤寒沙门氏菌鞭毛蛋白可增强DnaJ蛋白的免疫反应

摘要

To make sure the role of flagellin as mucosal adjuvants and the protective effect of streptococcus pneumonia infection when combined with DnaJ protein.Methods:Recombinant plasmid pET-28 ( a)/flagella was transferred to E.coli BL21(DE3).Over-expression of flagella was induced by IPTG and purification for animal study.All the C57BL/6 mice were randomly divided into three groups ,then respectively intranasally given the mixture of flagellin and DnaJ protein ( experimental group ) , DnaJ protein( control group 1 ) and the mixture of GST and DnaJ protein ( control group 2 ).The serum IgG and its subtype , cytokines secreted by mice spleen cells were detected by ELISA.At last all the C57BL/6 mice were intranasally immunized with Streptococcus pneumoniaD39.The protective effect by survival times were evaluated.Results: The mice of experimental group could secrete high level of serum IgG and cytokines IFN-γ,IL-4 and IL-17A.What more,the survival rate of mice in experimental group was 60%,a significant statistical difference with the control group.Conclusion:The flagellin as an adjuvant can reinforce the immune response of DnaJ protein and have better protection of resistance D 39 infection.We suggest that flagellin can be used as protein vaccine adjuvants.%目的：明确鞭毛蛋白的黏膜佐剂作用，并研究其与DnaJ蛋白联合免疫对肺炎链球菌感染的保护作用。方法：含有重组质粒pET-28（ a）/鞭毛蛋白基因的大肠杆菌DE3经IPTG诱导后表达鞭毛蛋白，纯化后用于后期动物实验如下：将C57BL/6小鼠随机分为四组通过鼻腔分别滴入鞭毛蛋白（10μg）与DnaJ蛋白（10μg）混合剂（实验组）、DnaJ蛋白（10μg）（对照组1）、DnaJ蛋白（10μg）与GST蛋白（10μg）混合剂（对照组2），每组均30μl。通过检测抗体的效价、亚型以及细胞因子水平来评估鞭毛蛋白能否提高肺炎链球菌DnaJ蛋白诱导的免疫效应；然后用肺炎链球菌D39（5×107 CFU）进行鼻腔攻毒，观察小鼠的生存率，以评估其保护效应。结果：实验组较对照组产生更高效价的血清IgG（其中主要产生IgG1）和更高水平的细胞因子IFN-γ、IL-17A和IL-4；另一方面实验组对致死性D39感染的保护率达60％，DnaJ蛋白免疫组的保护率为50％。结论：鞭毛蛋白与DnaJ蛋白联合应用可以增强宿主对DnaJ蛋白的免疫反应，并且对致死剂量的肺炎链球菌感染有保护作用，提示鞭毛蛋白可以作为肺炎链球菌DnaJ蛋白疫苗的佐剂应用。