Sj31与鼠IFN-γ基因融合DNA疫苗免疫保护效果的研究  

Studies on the protective immunity of DNA vaccine of rat IFN-γ infused with Schistosoma japaonicum tissue protein B.

作　　者：罗永慧[1] 易新元[2] 曾宪芳[2] 

机构地区：[1]江西省妇幼保健院检验科,江西南昌330008 [2]中南大学湘雅医学院寄生虫病研究室,湖南长沙410078

出　　处：《中国热带医学》2005年第7期1401-1404,1514,共5页China Tropical Medicine

基　　金：WHO/TDR资助项目(No.980268)

摘　　要：目的构建表达mIFN-γ与Sj31抗原融合蛋白的DNA疫苗,并探讨其免疫保护作用。方法在Sj31基因上游引物与mIFN-γ下游引物的5’分别设计与克隆载体相匹配的酶切位点,在mIFN-γ上游引物与Sj31基因下游引物的5’设计相同的酶切位点。分别进行PCR扩增,再通过引物的5’端相同的酶切位点进行2个PCR产物的酶切与连接,以连接产物为模板,应用Sj31的上游引物和mIFN-γ下游引物进行PCR扩增,将此次PCR产物经常规方法克隆入载体pcDNA3.0(简称为pc),构建成多价DNA疫苗pc-Sj31-mIFN-γ。对重组质粒鉴定后,大量制备纯化质粒免疫昆明小鼠,48只小鼠分为A、B、C、D4组,对照组的小鼠于股四头肌注射质粒pcDNA3.0100μg,3个免疫组的小鼠分别注射pc-Sj31、pc-mIFN-γ和pc-Sj31-mIFN-γ质粒DNA各100μg。在0、2、6周免疫共3次,第3次免疫后2周,每只小鼠经腹部贴片感染尾蚴40±1条,感染后第42天剖杀,计数各小鼠检获成虫数及肝卵数。结果pc-Sj31-mIFN-γ(D组)免疫组的减虫率和肝组织减卵率分别为28.56%和60.20%,但是表达鼠mIFN-γ与Sj31抗原融合蛋白的DNA疫苗并未明显优于单纯表达日本血吸虫组织蛋白酶B(Sj31)DNA疫苗的免疫保护作用。结论pc-Sj31-mIFN-γ多价DNA疫苗构建成功,但其诱导小鼠抗血吸虫病免疫保护效果不明显优于pc-Sj31。Objective To construct a fusion expression vector for DNA vaccine including murine interferon gamma （mIFN- γ ） and antigen Sj 31 to enhance the protective immunity of the Schistosoma japonicum cathepsin B（Sj 31 ）, and explore their immune protection against Sj . in mice. Methods Restriction enzyme sites that complemented with the eukaryotic expression plasmid pcDNA3.0 were respectively designed at the 5＇ end of Sj 31 gene＇s foreword primer and mIFN - y gene＇s reverse primer.The same restriction enzyme site was adopted in the design of the 5＇ end of mIFN - γ gene＇s foreword primer and Sj 31 gene＇s reverse primer. The two genes were amplified, the PCR products digested with the same enzyme for the 5＇ end of primer and llgated. The ligated product was PCR amplified with Sj 31 gene＇s foreword primer and mlFN - γ gene＇s reverse primer. The PCR product was then subcloned into the vector pcDNA3.0 （pc） by routine methods, pc - Sj 31 - mIFN - γ has been established. After verification, the antigen plasmids were intramuscularly inoculated to immunize the mice.48 mice were divided into four groups （A, B, C, D）. The mice in vaccination groups were immunized intramuscularly with 100 gg per mouse of each plasmid DNA, and those in control group were injected pcDNA3.0 plasmid DNA only at week 0,2,6. 2 weeks after the final injection, the mice were challenged percutaneously by 40 ± 1 cercariae, and were sacrificed 42 days after challenge. Worm burden and liver eggs were counted. Results Vaccination with pc - Sj 31 - mIFN - γ elitcited 28.56% of worm reduction rate and 60.20% of liver eggs reduction rate, And the worm and the liver eggs reduction rate in pc - Sj 31 - mIFN - γ DNA vaccine group was not signficant in comparision with pc - Sj 31 DNA vaccine group. Conclusion Construction of recombinant pc - Sj 31 - mIFN - γ was achieved. But it can not enhance the protective immunity elicited by pc - Sj 31 DNA vaccine in mice.

关 键 词：日本血吸虫 组织蛋白酶 B mIFN-γ 多价DNA疫苗 

分 类 号：R383.2[医药卫生—医学寄生虫学]