呼吸道融合病毒 ( Respiratory syncytia virus, RSV ) 已被認為是造成幼童與老年族群的下呼吸道感染的主因，此病毒不僅具高度傳染力還會不斷的在每年爆發大流行。病毒感染初期通常在人體呈現輕微感冒症狀，但隨著病毒感染時間延長有可能會發展為嚴重的下呼吸道疾病，例如：支氣管炎、肺炎等，而且截至目前為止仍無臨床上認可的疫苗。
根據對 RSV 先前的研究指出 RSV 表面的 fusion protein 可以引發中和性抗體，並且誘發細胞毒殺 ( CTL ) 反應、刺激 TH1 細胞分泌激素，例如： IL-2, IL-12, IFN-γ 以對抗 RSV 病毒的感染，更重要的是以 RSV fusion protein 做為抗原，不會產生疫苗反效加強疾病 ( vaccine-enhanced disease ) 的現象。
因此，我們選擇以完整的 RSV 融合蛋白 ( fusion protein, F ) 以及去除疏水性穿膜端 ( transmembrane domain truncated, F0ΔTM ) 兩種重組蛋白做為抗原，以此探討發展 RSV 疫苗的可能性。為了進行動物免疫實驗以研究融合蛋白 F0 / F0ΔTM 的抗原特性，我們選擇在大腸桿菌 BL21 (DE3) ，桿狀病毒昆蟲細胞 ( baculovirus / Sf9 insect cells ) ，以及哺乳類動物 293A 細胞 ( mammalian 293 cells )表達His-tagged 重組融合蛋白。這三種表現系統，都有各自不同的重組蛋白生產機制與轉譯後修飾能力。三種表現系統的表達產物皆以膠體電泳、西方墨點法、蛋白質胺端定序分析，根據結果我們發現哺乳類動物 293A 細胞所表現的融合蛋白 F0ΔTM 最符合實驗設計所需，因此我們將該重組蛋白純化後注射至小鼠，並與加熱去活化的 RSV B1 ( HI-RSV B1 ) 做比較，進行動物免疫反應分析；對小鼠血清進行抗體 IgG, IgG1, IgG2a 效價檢定、病毒毒力中和試驗。分析結果顯示：融合蛋白 F0ΔTM 所引起的抗病毒抗體效價與 HI-RSV 相同，而且抗體也具病毒中和能力。更重要的是，融合蛋白 F0ΔTM 誘發的 TH1 / TH2 免疫路徑偏向 TH1 。我們認為293A 細胞所表達的融合蛋白 F0ΔTM 是一個良好的保護性抗原，能誘發正確且強效的體液免疫。

Respiratory syncytia virus (RSV) has been recognized as the leading cause of lower respiratory infections in children and in elderly. Human RSV occurs in yearly outbreaks and is highly contagious. As a result, it causes mild cold symptoms following virus exposure could often progress to more serious lower respiratory tract diseases such as bronchilitis and pneumonia. Currently no effective vaccine is available.
Previous studies have shown that RSV fusion (F) protein elicits neutralizing antibody, induces CTL responses and stimulates an increased production of Th1 cytokines, IL-2, IL-12 and IFN-γ raised against live RSV infection and importantly does not induce vaccine-enhanced diseases.
In this rationality, full length RSV fusion protein (F) and transmembrane domain truncated F protein (F0ΔTM) are chosen as target antigens of RSV vaccine development. To investigate the antigenicity of F0 and F0ΔTM in animal immunization study, expression of His-tagged F and F0ΔTM in E. coli BL21 (DE3) cells, baculovirus/Sf9 insect cells, and mammalian 293 cells, which possess the different mechanisms of protein expression and translational modification, were performed. Recombinant protein expressed in three different systems were examined by performing SDS-PAGE, immunoblotting and peptide N-terminal sequencing. According to the results, recombinant fusion protein F0ΔTM expressed by mammalian 293 cells correspond to our original recombinant protein design. Therefore, this recombinant fusion protein F0ΔTM was purified and injected into BALB/C mice for animal immunization study compared with heat inactivated RSV B1 (HI-RSV B1). Mouse serum was performed with a series of assay including titer of antibody IgG, IgG1, IgG2a determination and plaque-reduction neutralization test. As the result, titer of anti-virus antibody raised by fusion protein F0ΔTM was same with HI-RSV B1 and also the ability of live RSV neutralization. Moreover, this antigen stimulates a immune response toward TH1 pathway. From the indications of these consequence, we could say that recombinant fusion protein F0ΔTM expressed by 293A cells is a good protective antigen for raising correct and strong humorial immunity.

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