以高壓流體注射法建立之小鼠模式探討B型肝炎病毒核心蛋白影響B型肝炎病毒清除之機制

Abstract

B型肝炎病毒在進入宿主後如何造成持續性感染是延宕已久的問題，之前利用以高壓流體力學注射方式活體轉染的小鼠模式來研究與病毒持續性表現或清除有關的病毒因子，已知B型肝炎核心蛋白是造成病毒清除的主要病毒因子，因此本篇論文的主要目的是進ㄧ步地探討其影響病毒清除的詳細機制。首先，利用一系列的核心蛋白突變株，我們發現核心蛋白第176到185個胺基酸片段為導致B型肝炎病毒清除的決定性區域，但是此區域並不包含重要的抗原決定位。當此片段缺失時會延長B型肝炎病毒在小鼠中的表現，但並不影響病毒在肝細胞中的複製、轉錄、與病毒抗原的表現。此外，此片段的缺失也會使小鼠對B型肝炎病毒無法產生適當的體液型及細胞型免疫反應。再者，利用核鞘組裝缺失的核心蛋白突變株，更進ㄧ步揭示核酸核鞘的結構對於核心蛋白導致病毒清除之必要性，同時包含在核酸核鞘中的病毒或是細胞之 RNA是影響免疫反應及病毒清除所必須。綜合上述的研究成果，我們提出了B型肝炎病毒核酸核鞘是B型肝炎病毒的病原相關分子模式的假說，這個病原相關分子模式與病毒清除與否息息相關。本研究對於這個目前仍在發展中的領域注入新的觀念，值得更深入地探究。

The mechanism of hepatitis B virus (HBV) persistence following its exposure has long been an unresolved question. Using a hydrodynamics-based mouse model for HBV persistence, we have previously found that HBV core protein (HBc) was the major determinant viral factor for HBV clearance. Therefore, my thesis aimed to further explore the underlying mechanisms. First, we determined the region of HBc protein that is crucial for HBV clearance in mice by using a series of HBcAg deletion mutants. Interestingly, the very C-terminal 10-residue region HBc176-185 of HBc protein, which does not appear to contain a major epitope of T and B cells, contributed to HBV clearance. A deletion of this region prolonged HBV persistence in mice while the viral replication, transcription and translation within hepatocytes remained unaffected. Besides, this particular HBV mutant failed to induce robust humoral and cellular immunity against HBV. Moreover, using an assembly-defective HBV mutant, HBcY132A pAAV/HBV1.2, we revealed the requirement of nucleocapsid structure for inducing effective immunity that leads to HBV clearance in mice. Viral or cellular RNAs, but not DNAs, within the capsid particle were essential for the nucleocapsid to trigger antiviral responses. Finally, these findings prompted us to hypothesize that the HBV nucleocapsid functions as a novel viral pathogen-associated molecular pattern (PAMP) that promotes the viral clearance. This is a burgeoning and exciting research area and may open a new field for future study.