細胞白介素-1β基因做為石斑虹彩病毒DNA疫苗佐劑之效果

虹彩病毒科裡共有五屬，其中細胞腫大病毒屬 (Megalocystovirus)對養殖漁業造成危害最為嚴重。本實驗是採用DNA疫苗來做為防治虹彩病毒的方法。先前以虹彩病毒錐體膜蛋白基因 (OSGIV-8L)作為抗原基因之DNA疫苗，另外導入一段點帶石斑魚細胞白介素1-β (Interleukin-1β, IL-1β)基因作為免疫佐劑，但並未執行攻毒試驗，因此本實驗測試IL-1β基因做為免疫佐劑的功效進而評估提升疫苗免疫效果。將本實驗DNA疫苗轉染 (Transfect)進石斑魚鰭細胞結果發現，細胞確實會表現出抗原蛋白。同樣的結果也在活體實驗中發現，接種DNA疫苗後石斑魚肌肉、脾臟在組織免疫化學染色 (Immunohistochemical, IHC）結果中發現皆有抗原被偵測到，其中又以含免疫佐劑IL-1β之DNA疫苗表現最多。而對於石斑魚免疫力增強部分，在實驗淋巴細胞吞噬能力、超氧陰離子產生立、溶菌酶活性中搭配IL-1β使用的DNA疫苗表現皆較控制組、不含免疫佐劑DNA組來的好。魚血清中專一性抗體的量亦是含免疫佐劑IL-1β之DNA疫苗組別最多。且IL-1β基因能誘導TNF-α、CD8α、IL-8之基因表現，進而在攻毒試驗中展現了良好的存活力 (90 %)。本實驗結果顯示，搭配免疫佐劑IL-1β基因之DNA疫苗能提升疫苗免疫效果。且IL-1β基因可能藉由提升石斑魚非特異性免疫、刺激免疫相關基因，進一步使DNA疫苗能更迅速低誘發特異性免疫反應，並使石斑魚對點帶石斑虹彩病毒產生抵抗力。這些結果顯示，細胞白介素1β作為DNA疫苗之基因免疫佐劑 (genetic adjuvants)的潛力。

關鍵字

點帶石斑魚；點帶石斑虹彩病毒； DNA疫苗；免疫佐劑；細胞白介素1β

並列摘要

Megalocytivirus is one of iridoviridae family and emerged in recent years as a troublesome pathogen to a wide range of marine and freshwater fish. Vaccination is one of the useful methods against viral disease. In previous study, we used a bicistronic vector encoding OSGIV-8L and grouper interleukin-1β (IL-1β) as a DNA vaccine. The expression of grouper IL-1β was applied to increase the efficiency of DNA vaccine. But, we didn't evaluate how much efficiency increased by IL-1β. So we aimed to determine the efficiency and function of IL-1β in DNA vaccine in this study. In the premiere results, the grouper fin cells were transfected with the DNA vaccine and the antigen protein were detected through western blot. Moreover, in the in vivo experiments, after vaccinating via intramuscular injection, OSGIV-8L's mRNA and protein were detected in spleen and muscle. Among the experimental conditions, the DNA vaccine containing IL-1β gene express more OSGIV-8L's mRNA and protein than the DNA vaccine without IL-1β gene. Furthermore, the immunological analysis showed that pIRES-8L/IL-1β was able to (Ⅰ) stimulate the respiratory burst of lymphocytes at 21 day post-vaccination, (Ⅱ) stimulate the phagocytic capacity of lymphocytes at 3day and 21day post-vaccination, (III) stimulate the lysozyme activity at 7day,14day and 21day post-vaccination, (IV) upregulate the expression of immune related genes, and (V) induce the production of serum antibodies. And when challenged groupers with 104 copies of OSGIV, the groupers vaccinated with pIRES-8L/IL-1β showed higher survive rate compared to control groupers and the groupers vaccinated with DNA vaccine without IL-1β gene. In summary, the DNA vaccine with IL-1β gene may enhance the grouper nonspecific immunity to induce specific immune response more rapidly, and develop resistance to virus. These results reveal that the potential application of fish IL-1β in fish vaccination.

並列關鍵字

Orange-spotted grouper ； Orange-spotted grouper iridovirus virus ； DNA vaccine ； immune adjuvant ； interleukin-1β

參考文獻

黃淑敏, 2010. 石斑魚虹彩病毒疫苗研發. 行政院農委會家畜衛生試驗所獸醫專訊. 3, 31-34.

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細胞白介素-1β基因做為石斑虹彩病毒DNA疫苗佐劑之效果

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