臺灣博碩士論文加值系統

廣東住血線蟲為大鼠肺寄生蟲，人類感染後常引發嗜伊性腦膜炎或嗜伊性腦膜腦炎，造成致命病變。目前尚未發展出有效的診斷及治療方法，對於在感染幼蟲後產生的免疫反應機制也缺乏完整性了解。本研究係以蛋白質體學分析廣東住血線蟲不同齡期蟲體蛋白質變化情形，初期目標係針對第三期及第五期幼蟲進行探討。比較第三期幼蟲與雌性第五期幼蟲，發現前者之蛋白質主要分布在分子量15-35 kDa、pI 值5-6 附近，後者主要分布在分子量40-100 kDa、pI 值5-7附近。以質譜分析119 來自廣東住血線蟲第三期幼蟲及雌性第五期幼蟲之差異性蛋白質點，再與NCBI 資料庫進行比對，共100 個蛋白質點被鑑定出來，33個來自第三期幼蟲，67來自第五期幼蟲。再與GO（gene ontology）資料庫比對，有35 個蛋白質點（第三期幼蟲6個，第五期幼蟲29 個）經分析後得知其功能性，依其功能分為12類，與壓力相關之蛋白質最多。推測廣東住血線蟲感染宿主後，在其主體內要適應新環境，亦要抵禦宿主之免疫反應，因此產生與壓力相關蛋白質來抵抗環境。

Angiostrongylus cantonensis is a parasite of rats. It is an important causative agent of eosinophilic meningitis and eosinophilic menigoencephalisis in humans. This study was conducted to investigate the alternations of proteomes on the third- and fifth-stage larvae of A.cantonensis by proteomic techniques. Proteins with molecular weight of
15-35 kDa and pI 5-6 were mainly revealed in the third-stage larvae whereas those of 40-100 kDa and pI 5-7 in the female fifth-stage larvae. Of 119 protein spots identified by MALDI-TOF MS and database searches, 100 were found to have significant similarities with existing proteins in the NCBI database: 33 from the third-stage larvae and 67 from
the fifth-stage larvae. After matching with the GO (gene ontology) database, the functions of 35 spots (6 from the third-stage larvae and 29 from the fifth-stage larvae) were determined. According to their functions, these spots were classified into 12 groups and stress-related proteins were
found to be the most abundant. The high expression level of stress-related proteins reflects that the larvae of A. cantonensis need to adapt the new environment and the resist the host immune responses after entering the
new host.

目錄
指導教授推薦書‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥ii
論文口試委員會審定書‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥iii
長庚大學授權書‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥iv
致謝‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥v
中文摘要‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥vi
英文摘要‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥vii
目錄‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥viii
圖表目錄‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥ix
緒論‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥1
材料與方法‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥11
結果‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥18
討論‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥22
參考文獻‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥32
附圖及表格‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥40
附錄‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥69

圖表目錄
圖一、廣東住血線蟲的分佈及全球流行的區域。‥‥‥‥‥‥‥‥40
圖二、廣東住血線蟲生活史。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥41
圖三、2006-2007 間，中國地區軟體動物感染廣東住血線蟲之流行病
學調查。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥42
圖四、人類廣東住血線蟲症的相關症狀。‥‥‥‥‥‥‥‥‥‥‥43
圖五、在解剖顯微鏡下觀察自扁捲螺收集之廣東住血線蟲第三期幼蟲
形態。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥44
圖六、在解剖顯微鏡下觀察自感染後23 天之大鼠腦部收集之廣東住
血線蟲第五期幼蟲形態。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥45
圖七、自感染後50 天之大鼠右心室及肺動脈收集之廣東住血線蟲成
蟲。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥46
圖八、實驗室建立之廣東住血線蟲生活史。‥‥‥‥‥‥‥‥‥‥47
圖九、以聚丙烯醯胺膠體電泳（SDS-PAGE）分析廣東住血線蟲第五
期幼蟲（雌、雄）蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥48
圖十、以pH 3-10 的膠條進行二維蛋白質電泳，分析廣東住血線蟲雄
性成蟲蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥49
圖十一、以pH 3-10 的膠條進行二維蛋白質電泳，分析廣東住血線蟲
x
雌性成蟲蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥50
圖十二、以pH 3-10 的膠條進行二維蛋白質電泳，分析廣東住血線蟲
雄性第五期幼蟲蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥51
圖十三、以pH 3-10 的膠條進行二維蛋白質電泳，分析廣東住血線蟲
雌性第五期幼蟲蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥52
圖十四、以pH 4-7 的膠條進行二維蛋白質電泳，分析廣東住血線蟲
雄性第五期幼蟲蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥53
圖十五、以pH 4-7 的膠條進行二維蛋白質電泳，分析廣東住血線蟲
第五期幼蟲雌蟲蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥54
圖十六、以pH 4-7 的膠條進行二維蛋白質電泳，分析廣東住血線蟲
第三期幼蟲蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥‥‥55
圖十七、以pH 4-7 的膠條進行二維蛋白質電泳，分析廣東住血線蟲
雄性成蟲蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥56
圖十八、以pH 4-7 的膠條進行二維蛋白質電泳，分析廣東住血線蟲
雌性成蟲蛋白質組成。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥57
圖十九、比對廣東住血線蟲第三期及雌性第五期幼蟲在二維蛋白質電
泳膠體上蛋白質分佈的差異。‥‥‥‥‥‥‥‥‥‥‥‥58
圖二十、廣東住血線蟲雌性第五期幼蟲差異性蛋白質點。‥‥‥59
圖二十一、廣東住血線蟲第三期幼蟲差異性蛋白質點。‥‥‥‥60
xi
圖二十二、利用MASCOT資料庫搜尋引擎把差異性蛋白質點和廣東
住血線蟲表現序列標幟資料庫相互比對。‥‥‥‥‥61
圖二十三、NCBI資料庫比對到之蛋白質點與GO資料庫進行比
對。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥62
圖二十四、與GO(gene ontology)比對到之蛋白質點依其功能性進行
分類。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥63
圖二十五、截取出與GO資料庫比對到之蛋白質點。‥‥‥‥‥‥64
圖二十六、壓力相關之蛋白質點，依其功能分成三大類。‥‥‥65
圖二十七、以LC-MS/MS質譜儀分析廣東住血線蟲第五期幼蟲整體
蛋白質之表現。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥66
表一、人類廣東住血線蟲症案例及發生國家。‥‥‥‥‥‥‥‥‥67
表二、與GO資料庫比對到之蛋白質點，列出與壓力相關之蛋白
質。‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥‥68

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