登革病毒 (dengue virus; DENV) 是一種藉由蚊子所傳播的病毒，病人感染登革病毒後主要出現的臨床症狀有從較輕微的登革熱 (dengue fever; DF) 至嚴重的登革出血熱 (dengue hemorrhagic fever; DHF) 及登革休克症 (dengue shock syndrome; DSS)。疾病盛行的區域主要為熱帶以及亞熱帶地區。根據WHO的統計，每年至少有五十至一百萬登革感染的案例發生，然而現今仍未有疫苗可供使用。實驗室先前的研究發現非結構性蛋白1 (NS1)蛋白質之C端序列與自體抗原具有高度相似性，因此抗登革病毒NS1之抗體會與人類的內皮細胞以及血小板產生交互作用，導致內皮細胞凋亡以及血小板凝集機制異常。考量以NS1發展疫苗的安全性，我們將具有交互作用的C端抗原片段進行修飾，產生了名為DJ NS1的嵌合蛋白。先前研究證實DJ NS1抗體對人類內皮細胞及血小板的黏附能力較全長DENV NS1抗體低，另外在小鼠模式中也顯示，給予小鼠DJ NS1抗體的治療可以有效的減緩DENV感染後所造成出血時間延長的現象，證實抗DJ NS1抗體具有保護小鼠對抗DENV感染之效果。為了進一步探討在主動免疫小鼠模式中，DJ NS1蛋白是否也能提供小鼠對抗DENV感染之保護效果，我們利用高分子型奈米複合物以及鋁鹽作為佐劑，對小鼠進行兩次接種。實驗結果發現，利用高分子型奈米複合物作為佐劑，與鋁鹽相較能引起小鼠體內更高效價的抗體反應，而且依據IgG isotypes顯示會產生Th1和Th2免疫反應。另外，在in vitro 或是in vivo的細胞毒性測試實驗中，我們發現不管是給予包裹DJ NS1的高分子型奈米複合物或是單獨給予高分子型奈米複合物對於THP-1細胞或是小鼠都不具有毒性，顯現其安全性。為了進一步研究包裹DJ NS1蛋白之高分子型奈米複合物引起之主動免疫保護效果，小鼠在接種包裹DJ NS1的高分子型奈米複合物後，給予高劑量的登革病毒進行感染。實驗結果發現，接種包裹DJ NS1之高分子型奈米複合物的小鼠，可以有效減緩因感染DENV後所造成出血時間延長的現象、病毒抗原表現量、以及免疫細胞浸潤。除此之外，我們也更進一步發現包裹DJ NS1蛋白之高分子型奈米複合物在接種小鼠後，可以引發持久的抗體反應並且提供小鼠長期的保護效果。雖然包裹DJ NS1的高分子型奈米複合物如何提供小鼠保護效果的詳細機制仍然有待進一步探討，但是綜合以上實驗結果，證明了利用高分子型奈米複合物包裹DJ NS1蛋白可以有效的引發小鼠主動免疫反應，並且提供小鼠對抗登革病毒之保護效果。重要的是，實驗結果顯示包裹DJ NS1蛋白之高分子型奈米複合物提供了具潛力的登革疫苗策略。

Dengue virus (DENV) is an arthropod-borne virus which causes diseases ranging from dengue fever to dengue hemorrhagic fever and dengue shock syndrome in tropical and subtropical regions of the world. The WHO estimates that there are approximately 500,000-1,000,000 dengue cases annually, but there is no licensed dengue vaccine available. We previously showed that antibodies (Abs) directed against DENV nonstructural protein 1 (NS1) could cross-react with host cells due to the C-terminal region of DENV NS1 contains homologous sequences. Therefore, we generated a C-terminal modified DJ NS1 as a potential candidate for dengue vaccine. The endothelial cell and platelet binding activity of anti-DJ NS1 was lower than that of anti-DENV NS1. In addition, anti-DJ NS1 Abs reduced DENV-induced mouse tail prolonged bleeding time, showing that anti-DJ NS1 Abs may provide protection against DENV infection. In this study, we use polymer-based nanocomplexes or alum as an adjuvant to examine the protective effects of DJ NS1 in DENV-induced hemorrhage mouse model. Polymer-based nanocomplexes provide better adjuvant activity than alum and can induce both Th1 and Th2 responses as indicated by the production of specific Abs as well as IgG isotypes. In in vitro and in vivo cytotoxicity tests, there was no toxicity of DJ NS1-encapsulated nanocomplexes or nanocomplexes alone in THP-1 cells and in mice. Active immunization with DJ NS1-encapsulated nanocomplexes significantly reduced the DENV-induced prolonged bleeding time, viral antigen expression and macrophage infiltration to local infection sites. Furthermore, DJ NS1-encapsulated nanocomplexes induced longer Ab persistence than alum and provided long-term protection. The detailed mechanisms of the protective effect remain to be further investigated. Taken together, DJ NS1-encapsulated nanocomplexes induce effective immune responses and provide protection against DENV infection. Importantly, the results provide support for new strategies for the development of protective dengue vaccines.

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