【作者】 常茂林； 叶惟虎； 李宇红； 樊明文；

【Author】 Maolin Chang;Weihu Ye;Yuhong Li;Mingwen Fan;The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) & Key Laboratory of Oral Biomedicine of Ministry of Education(KLOBM),School & Hospital of Stomatology,Wuhan University;

【机构】 武汉大学口腔医学院；

【摘要】 目的:比较三种不同类型的免疫佐剂对防龋亚单位疫苗黏膜免疫增强效应及其对免疫反应类型的影响。方法:分别采用鞭毛蛋白、AddaVax及PIKA佐剂联合防龋亚单位疫苗PAc以不同给药途径免疫小鼠。并在免疫前及免疫后2、4、6、8、10、12、16及20周收集小鼠的唾液和血液样本,应用ELISA技术检测样本中特异性抗PAc IgA和抗PAc IgG抗体的浓度,并测定IgG1和IgG2a水平分析IgG1/IgG2a比值。于末次免疫后8周处死小鼠,分离脾脏,经脾细胞培养和Pac刺激后,提取脾细胞总RNA,RT-qPCR测定检测IL-4及IFN-γmRNA水平,收集脾细胞上清,检测脾细胞分泌的IL-4及IFN-γ的水平。结果:与单纯的疫苗(Pac免疫组)对照组相比较,三种佐剂均可提高小鼠血清和唾液特异性抗Pac抗体水平,各免疫组第0周时(接种疫苗前)血清IgG1/IgG2a较低,自免疫后第2周呈起逐渐升高。PIKA+PAc和鞭毛蛋白+PAc滴鼻组唾液抗PAc IgA水平在第4,6,8周均显著高于其他各组(P<0.05,P<0.05,P<0.01)。经PAc刺激48h后,脾细胞IL-4 mRNA和IFN-γmRNA水平在各实验组之间无统计学差异(P>0.05)。各实验组的脾细胞培养上清中IL-4水平均高于阴性对照组(空白未刺激组)(P<0.05),所有实验组及对照组脾细胞上清的IFN-γ水平之间无统计学差异(P>0.05)。结论:不同佐剂分别与防龋亚单位疫苗PAc共同经滴鼻免疫小鼠后所诱导的免疫效应的影响不一致,其中PIKA、鞭毛蛋白对防龋亚单位疫苗PAc有较好的黏膜佐剂效应,可有效提升小鼠唾液特异性IgA抗体水平。更多还原

【Abstract】 Objective:The aim of the study was to compare the efficacy of anti-caries subunit vaccine rPAc with flagellin,AddaVax or PIKA as an adjuvant by intranasal immunization,respectively.Methods:Anti-caries subunit vaccine rPAc with flagellin,AddaVax or PIKA as an adjuvant,respectively,was delivered into mice by intranasal immunization.Saliva and serum were collected at 0,2,4,6,8,10,12,16,20 weeks and the specific anti-caries IgA and IgG antibody were analyzed by using ELISA method.The specific subclass of anti-PAc IgG in serum was also measured.Mice were executed 8 weeks after the last immunization,and splenic cells were collected,the level of IL-4 and IFN-γ of splenic cells was measured by RT-PCR and ELISA.Result:Flagellin,AddaVax or PIKA can increase the level of anti-Pac IgG and salivary IgA,Among all groups,the concentration of anti-caries specific IgA of PIKA+ PAc group was higher than any other groups at the 4th,6th and 8th week.At the 6th week,the serum specific anti-PAc IgG in all groups reached the peak concentration,The IgG1/IgG2 a ratio was relatively low at the 0th week,and it rised gradually since the2 nd week,and the IL-4 concentration in PAc+PIKA and PAc+ Flagellin(i.n.) group was higher than that of other group.Conclusion:The anti-caries vaccine rPAc with flagellin or PIKA showed good immune efficacy in mice when intranasally immunized.更多还原