DNA Vaccines against Dengue Virus Type 2 Based on Truncate Envelope Protein or Its Domain III
DNA Vaccines against Dengue Virus Type 2 Based on Truncate Envelope Protein or Its Domain III
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DOI：
10.1371/journal.pone.0020528
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发表时间：
2011-07-11
期刊：
影响因子：
3.7
通讯作者：
Alves, Ada M. B.
中科院分区：
文献类型：
--
作者：
Azevedo, Adriana S.;Yamamura, Anna M. Y.;Alves, Ada M. B.
构建了两种DNA疫苗，其编码与人组织纤溶酶原激活物信号肽（t-PA）融合的DENV 2包膜蛋白（pE 1D 2）的胞外域（结构域I、II和III）或仅编码其结构域III（pE 2D 2）。重组蛋白的表达和分泌在体外证实在BHK细胞转染的两个质粒，检测代谢标记的基因产物的免疫荧光或免疫沉淀，使用多克隆和单克隆抗体DENV 2。此外，结果表明，E蛋白的胞外域可以在体内有效地表达，在哺乳动物系统中，没有prM蛋白，假设在登革热感染期间作为伴侣蛋白。用DNA疫苗免疫Balb/c小鼠，并用致死剂量的DENV 2进行攻击。所有pE 1D 2免疫的小鼠在攻击后存活，而45%的pE 2D 2免疫的动物在感染后死亡。此外，pE 1D 2免疫的小鼠中只有10%在攻击后表现出一些感染的临床症状，而用pE 2D 2接种的大多数动物表现出具有高发病率的疾病效应。pE 1D 2免疫小鼠的中和抗体水平显著高于pE 2D 2免疫动物，这也表明pE 1D 2疫苗比pE 2D 2更具保护性。
Two DNA vaccines were constructed encoding the ectodomain (domains I, II and III) of the DENV2 envelope protein (pE1D2) or only its domain III (pE2D2), fused to the human tissue plasminogen activator signal peptide (t-PA). The expression and secretion of recombinant proteins was confirmed in vitro in BHK cells transfected with the two plasmids, detected by immunofluorescence or immunoprecipitation of metabolically labeled gene products, using polyclonal and monoclonal antibodies against DENV2. Besides, results reveal that the ectodomain of the E protein can be efficiently expressed in vivo, in a mammalian system, without the prM protein that is hypothesized to act as a chaperonin during dengue infection. Balb/c mice were immunized with the DNA vaccines and challenged with a lethal dose of DENV2. All pE1D2-vaccinated mice survived challenge, while 45% of animals immunized with the pE2D2 died after infection. Furthermore, only 10% of pE1D2-immunized mice presented some clinical signs of infection after challenge, whereas most of animals inoculated with the pE2D2 showed effects of the disease with high morbidity degrees. Levels of neutralizing antibodies were significantly higher in pE1D2-vaccinated mice than in pE2D2-immunized animals, also suggesting that the pE1D2 vaccine was more protective than the pE2D2.