抗伯氏疟原虫GAP40蛋白免疫血清对动合子发育抑制作用的研究  

INHIBITIVE EFFECTS ON GLIDEOSOME-ASSOCIATED PROTEIN 40 OF PLASMODIUM BERGHEI ON ITS OOKINETES DEVELOPMENT

作　　者：何璐 朱俐颖 李思琦 陈露萌 曹雅明 朱晓彤 HE Lu;ZHU Li-Ying;LI Si-Qi;CHEN Lu-Meng;CAO Ya-Ming;ZHU Xiao-Tong(Department of Immunology,College of Basic Medical Science,China Medical University,Shenyang 110122,Liaoning,China)

机构地区：[1]中国医科大学基础医学院免疫学教研室,沈阳110122

出　　处：《寄生虫与医学昆虫学报》2021年第4期207-213,236,共8页Acta Parasitologica et Medica Entomologica Sinica

基　　金：国家自然科学基金面上项目(No.81971961)。

摘　　要：为探讨伯氏疟原虫滑动体相关蛋白40(P.berghei glideosome-associated protein 40,PbGAP40)作为传播阻断疫苗候选抗原的可行性,PCR扩增GAP40^(329-458 aa)编码基因并克隆入pET32a(+)载体,IPTG诱导PbGAP40重组蛋白(rPbGAP40)表达,纯化后免疫BALB/c小鼠。采集抗-PbGAP40免疫血清,通过ELISA和Western blot方法检测抗-PbGAP40免疫血清抗体滴度和特异性,IFA检测PbGAP40蛋白在伯氏疟原虫动合子期定位情况,体外实验检测抗-PbGAP40免疫血清对动合子期原虫发育的抑制效果。结果显示,成功表达rPbGAP40,抗-PbGAP40免疫血清抗体滴度为1∶51200;PbGAP40在合子、retort和成熟动合子表面表达。与对照组相比,抗-PbGAP40免疫血清处理组动合子生成数目下降36.5%(P<0.05),动合子转化率下降8.5%(P<0.05)。体外培养24 h后,抗-PbGAP40免疫血清处理组中61.2%的疟原虫停留在"retort"发育阶段。结果表明,PbGAP40可作为传播阻断疫苗的候选抗原。To characterize Plasmodium berghei glideosome-associated protein 40(PbGAP40)as a transmission blocking vaccine candidate,an epitope(329-458 aa)of GAP40 was amplified by PCR and ligated into pET32 a(+)plasmid,the recombinant PbGAP40 protein(rPbGAP40)was induced by IPTG,purified and immunized to BALB/c mice.Then,the anti-PbGAP40 immune sera were collected and purified.The specificity and antibody titer of the anti-PbGAP40 sera were determined by ELISA and Western Blot,respectively.The localization of PbGAP40 protein in ookinetes was demonstrated by IFA.The transmission blocking activity of anti-PbGAP40 sera on ookinete formation and differentiation was determined in vitro.In this study,we successfully induced rPbGAP40 expression and the antibody titer of anti-PbGAP40 sera was screened around 1∶51200.PbGAP40 was shown expressed on the surface of zygotes,retorts and matured ookinetes.Compared to the control groups,anti-PbGAP40 sera supplements reduced the number and conversion rate of ookinete by 36.5%and 8.5%,respectively(P<0.05).Furthermore,after 24 hours culture in vitro,61.2%of the anti-PbGAP40 sera treatedstayed as“retort”stage.Together,PbGAP40 could be utilized asaoptional transmission blocking vaccine candidate.

关 键 词：伯氏疟原虫 滑动体 有性发育 滑动体相关蛋白GAP40 

分 类 号：R392[医药卫生—免疫学]