2014-01-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/649319摘要：高病原性禽流感 H5N1 病毒能造成人類高致死率的呼吸道感染疾病，開發能引發黏膜性免疫反應的 H5N1 疫苗為重要課題。本計畫依據類鐸受體配體可作為疫苗佐劑，增強全身性和黏膜性免疫反應的方式，以能活化 TLR-2/1 反應的第二 b 型忌熱性腸毒素 B次單元 (LTIIb-B5)作為黏膜佐劑，進一步研發 H5N1 黏膜疫苗。研究時將對於 LTIIb-B5蛋白 upper pore 區 Met69, Ala70, Leu73, and Ser74 與 lower pore 區 Thr13 胺基酸突變，評估其對 TLR-2/1 活化效果和對神經節苷脂 GD1a 結合的影響。接著，LTIIb-B5突變蛋白會與 H5HA 重組抗原混合，以鼻腔滴注方式引發 BALB/c 小鼠免疫反應，並測試其黏膜性與全身性免疫反應以及攻毒後保護力。最後，我們會以 sortase A 蛋白酶將 H5HA 與LTIIb-B5 結合成融合抗原，以改進此黏膜疫苗之效能。本研究成果可提供發展 H5N1 黏膜疫苗的有用資訊。<br> Abstract: Highly pathogenic avian influenza (HPAI) H5N1 viruses can cause severe respiratory diseases in humans with a relatively high mortality rate. Development of H5N1 mucosal vaccines is thus particularly interesting for generating protective mucosal immunity. Since Toll-like receptor (TLR) ligands can act as both the systematic and mucosal adjuvants, we propose to investigate the type IIb heat labile enterotoxin B subunit (LTIIb-B5), a TLR-2/1 ligand as the mucosal adjuvants for H5N1 vaccine development. Site-directed mutagenesis studies will be performed at the residues Met69, Ala70, Leu73, and Ser74 in the upper core region and at the residue Thr13 in the lower pore region of the LTIIb-B5 pentamer to investigate their effects on TLR-2/1 activation and GD1a ganglioside binding. BALB/c mice will be intranasal immunized by H5HA recombinant protein plus LTIIb-B5 mucosal adjuvant to evaluate mucosal and systemic immune responses as well as protective immunity against live virus challenge. Direct linkage of LTIIb-B5 to H5HA antigen will be assessed using sortase A-mediated posttranslational fusion to further improve the mucosal vaccine adjuvanticity. Our findings may provide useful information for developing mucosal H5N1 vaccines.禽流感疫苗黏膜性免疫力LTIIb-B5第 2/1 型類鐸受體GD1a 神經節苷脂avian influenza vaccinemucosal immunityLTIIb-B5TLR2/1GD1a ganglioside