- 學位論文
人類 SCARB2蛋白對於腸病毒感染的機制和動物模式的建立
Establishment of human SCARB2 receptor transgenic mouse model for enterovirus 71 infection
摘要
腸病毒71型（EV71）是一種感染幼兒後主要會引起手足口症的病毒，在較嚴重的病例中，甚至會引發神經病變(例如腦膜炎)進而導致高死亡率的發生。及至今日，EV71疫情爆發主要分布在東南亞及南亞地區，包括了台灣、馬來西亞、新加坡、日本和中國大陸等。然而，以公共衛生宣導來控制腸病毒傳播的成效仍顯不足。因此，研究EV71感染細胞的途徑以及其誘發的病理症狀，包括手足口症和神經病變症狀，藉以發展出有效的治療型藥物或是預防型的疫苗，來控制腸病毒疫情至關重要。最近有兩篇關於腸病毒受器的論文發表，其一是人類scavenger receptor class B, member 2 (human SCARB2, hSCARB2)，它在全身多數的體細胞都有表現；其二是P-selectin glycoprotein ligand-1 (PSGL-1)，主要表現在白血球上。因此，我們選擇了hSCARB2做為研究的主軸。為了要找出EV71感染細胞的機轉，我們建構了有表現hSCARB2 蛋白的老鼠纖維母細胞株NIH3T3-SCARB2細胞，結果發現腸病毒藉由hSCARB2感染宿主細胞主要是由clathrin和dynamin調控的胞吞作用，且caveolae並未參與。接下來為了要找出hSCARB2在EV71感染宿主中扮演的角色，我們建構了表現hSCARB2的基因轉殖鼠（hSCARB2-Tg mice）進行EV71感染試驗。結果發現此種基因轉殖鼠的確能夠被EV71感染。以B4或B5基因型的EV71感染基因轉殖鼠發生的症狀類似於人類手足口症，且部分帶有輕微的神經病變症狀；而嚴重的神經病變導致跛足、甚至死亡可以在C2、C4和克沙奇16型感染此基因轉殖鼠後觀察到。病毒量和發炎相關細胞激素與EV71誘發的症狀嚴重度有高度相關。同時，我們也將此動物模式應用在測試EV71中和性抗體N3和潛在抗EV71藥物17-AAG對EV71感染後的保護效力。結果顯示兩者均能有效降低EV71感染後引發之病理症狀及提高存活率。根據以上結果可知hSCARB2-Tg mice是一種可被EV71感染及誘發相關病症，並能應用於觀察及研究的可信賴的動物模式。瞭解了EV71的感染機制和感染動物模式的建立後，除了可根據EV71感染機制研發治療藥物，且動物模式可成為評估EV71疫苗效力和治療藥物效果的測試平台，期望能夠加速對抗腸病毒的醫療發展腳步，使更多孩童受惠
關鍵字
腸病毒71型並列摘要
Enterovirus 71 (EV-71) is a caustic agent for hand, foot, and mouth disease (HFMD) in young children, but in some severe cases, neurological disorders with high mortality rates had been found. To date, epidemic outbreaks of EV71 have been reported mainly in Southeast and East Asia including Taiwan, Malaysia, Singapore, Japan, and China. The control of EV71 infection through public health interventions remains ineffectual and the treatments are unmet. Thus, study of the mechanisms of EV71 entry and it’s pathogenesis including HFMD and central nerve system (CNS) syndromes to develop a useful effective medications or prophylactic vaccines is important. Recently two EV71 receptors, human scavenger receptor class B, member 2 (SCARB2) expressed in the most of human tissues and P-selectin glycoprotein ligand-1 (PSGL-1) which restrictively expressed in the leukocytes, had been identified. Therefore, we chose human SCARB2 as a target for my research. In order to study the mechanism of EV71 infection, we made a human-SCARB2-transgenic mice fibroblast cell line, NIH3T3-SCARB2 cells, to investigate the pathway of EV71 entry. We had demonstrated that the EV71 entry into its host cells through a clathrin and dynamin-mediated but not caveolae-mediated endocytosis pathway. This pathway is a human hSCARB2-dependence. Following to understand the role of human SCARB2 in EV71-mediated diseases, we established a transgenic animal expressing human SCARB2. We found that this transgenic animal (hSCARB2-Tg mice) was highly susceptible to natural EV71 infection. The diseases observed in B4 or B5 genotype of EV71-infected hSCARB2-Tg mice was human-like HFMD syndromes and CNS dysfunction. Severe to death of CNS-like limb paralysis was observed in C2 and C4 of EV71 and even CVA16 infected hSCARB2-Tg mice. Viral loads associated with proinflammatory chemokines secretion were correlated with EV71-mediated pathogenesis. Also, we applied this animal model to evaluate the protective efficacy of EV71 neutralizing antibody N3 and potential anti-viral drug 17-AAG. Both of them can protect mice from EV71-induced illness and survive. These results implied that hSCARB2-Tg mouse is a reliable model for studying the EV71 pathogenicity in vivo. Understanding of the mechanisms of EV71 infections as well as infectious model establishment provide evidences to accelerate anti-EV71 medications development and serve as a platform to evaluate the efficacy of the vaccine candidates and antiviral drugs.
參考文獻
延伸閱讀
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