日本血吸虫体表蛋白Tetraspanin 2-A核酸疫苗的构建及其对小鼠的免疫试验

采用PCR法扩增日本血吸虫体表四跨膜家族蛋白2-A（SjTsp2-A）基因，构建重组质粒pcDNA3.1（+）/SjTsp2-A，将其转至大肠埃希菌DH5α制备DNA疫苗pcDNA3.1（+）/SjTsp2-A。24只BALB/c小鼠均分3组，每鼠于左股四头肌注射0.5 mg/ml盐酸布比卡因50 μl。次日，A组同法注射DNA疫苗pcDNA3.1（+）/SjTsp2-A，B组注射重组质粒pcDNA3.1（+）/SjGST，C组注射空质粒pcDNA3.1（+）。注射剂量均为100 μg/只。每隔2周注射1次，共3次，末次免疫后2周各组均经腹部皮下感染日本血吸虫尾蚴40±2条/鼠，45 d后剖杀，计数减虫率和减卵率。ELISA检测抗体效价，A组化分析股四头肌局部组织蛋白表达情况。结果A组的平均检虫数和每克肝组织虫卵数均显著低于B组和C组（P值均<0.05），A组的减虫率和减卵率分别为44.4%和28.4%。A组血清抗体效价高达1 ∶ 25 600。A、B两组局部组织均有特异性蛋白表达。DNA候选疫苗pcDNA3.1（+）/SjTsp2-A能诱导小鼠产生一定的免疫保护作用。

Construction of DNA Vaccine pcDNA3.1（＋）/Tetraspanin 2-A against Schistosoma japonicum and its Immuno-protective Effect in Mice

Tetraspanin 2-A （SjTsp2-A） gene was amplified by PCR. pcDNA3.1（+）/SjTsp2-A recombinant plasmids were constructed and transformed into E. coli DH5α. Twenty four BALB/c mice were randomly divided into pcDNA3.1（+）/SjTsp2-A group （A）, pcDNA3.1（+）/SjGST group （B） and pcDNA3.1（+） group （C）. Each mouse was injected through musculus quadriceps femoris by three times （two weeks interval） respectively with 100 μg pcDNA3.1（+）/SjTsp2-A, pcDNA3.1（+）/SjGST, or pcDNA3.1（+）. At two weeks after the final inoculation, mice were each challenged by 40±2 cercariae of S. japonicum. Forty-five days after infection， all mice were sacrificed, the number of worms collected and eggs in liver tisssue was counted. Anti-pcDNA3.1（+）/SjTsp2-A antibody was detected by ELISA and protein expression in quadriceps muscle by immunohistochemical staining. The worm reduction rate （44.4%） and egg reduction rate （28.4%） of group A was higher than those of group B and C （P＜0.05）, but no significant difference between groups B （3.9%, 19.3%） and C. Higher antibody titer （1 ∶ 25 600） was detected in sera of group A. Immunohistochemistry analysis showed an expression of specific antigens in quadriceps muscles of groups A and B. The DNA candidate vaccine induces partial protective immunity against S. japonicum in BALB/c mice.