基于壳聚糖及其衍生物的鱼用口服疫苗佐剂体系的研究  

Study of a Fish Oral Vaccine Adjuvant System Based on Chitosan and Its Derivatives

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作  者:刘晓璇 苗春宇 吴楠 奥浩之 吴颉[2] LIU Xiaoxuan;MIAO Chunyu;WU Nan;OKU Hiroyuki;WU Jie(Division of Molecular Science,Faculty of Science and Technology,Gunma University,Ota,Gunma 373-0057,Japan;State Key Laboratory of Biochemical Engineering,Key Laboratory of Biopharmaceutical Preparation and Delivery,Institute of Process Engineering,Chinese Academy of Sciences,Beijing 100190,China)

机构地区:[1]群马大学理工学部分子科学部,太田373-0057 [2]中国科学院过程工程研究所生化工程国家重点实验室,北京100190

出  处:《中国生物工程杂志》2024年第10期10-17,共8页China Biotechnology

基  金:国家自然科学基金(22078335)资助项目。

摘  要:目的:优化基于壳聚糖及其衍生物的鱼用疫苗佐剂的制备工艺、配方和评价方法。方法:以特定分子量和浓度的壳聚糖为原料,使用适合大规模生产的高压均质和喷雾干燥法制备壳聚糖及其季铵盐颗粒佐剂,负载灭活鳗弧菌后与鱼油均质乳化制备成口服疫苗,与饲料混合后投喂大菱鲆;调整鳗弧菌浓度和浸泡时间,探索建立适配的浸泡攻毒模型;引入海藻酸钠、β-葡聚糖等多种成分与壳聚糖及其衍生物组成复合佐剂,利用浸泡攻毒模型,通过攻毒后大菱鲆的存活率与免疫相关基因表达水平评价不同复合佐剂的免疫保护效果。结果:使用分子量50 kDa、质量浓度10 mg/mL的壳聚糖及壳聚糖季铵盐制备得到粒径220 nm的均一的颗粒作为口服疫苗佐剂;建立了用10~8 CFU/mL鳗弧菌浸泡12 h的大菱鲆浸泡攻毒模型;在浸泡攻毒实验中,壳聚糖季铵盐颗粒与海藻酸钠的复合佐剂配方能够使MHC-Ⅰ等免疫相关细胞因子的表达水平增加2~8倍,使大菱鲆存活率从0%提高至30%。结论:使用了适合放大生产的制备工艺得到壳聚糖及其季铵盐颗粒佐剂,在鳗弧菌浸泡攻毒实验中,壳聚糖季铵盐颗粒与海藻酸钠的复合佐剂有效增强了口服疫苗对大菱鲆的免疫保护,为基于壳聚糖及其衍生物的鱼用口服疫苗佐剂体系的开发与工业化生产提供了基础。Objective:The aim of this study is to optimize the preparation process,formulation,and evaluation methods for fish vaccine adjuvants derived from chitosan and its derivatives.Methods:Chitosan and its quaternary ammonium salt granule adjuvant were synthesized from chitosan of specific molecular weight and concentration through high-pressure homogenization and spray-drying techniques suitable for large-scale production.These adjuvants were loaded with inactivated Vibrio anguillarum,subsequently homogenized and emulsified with fish oil to produce an oral vaccine,which was incorporated into the feed and administered to turbot.The concentration of Vibrio anguillarum and the duration of immersion were systematically adjusted to establish an optimal immersion challenge model.Additionally,sodium alginate,β-glucan,and chitosan derivatives were combined to form a composite adjuvant.The immunoprotective efficacy of various composite adjuvants was assessed by evaluating the survival rates and the expression levels of immunity-related genes in turbot after exposure using the immersion challenge model.Results:Chitosan and chitosan quaternary ammonium salt,characterized by a molecular weight of 50 kDa and a mass concentration of 10 mg/mL,were utilized to generate homogeneous particles with a particle size of 220 nm,which served as oral vaccine adjuvants.An immersion challenge model was established for turbot where fish were exposed to Vibrio anguillarum at a concentration of 10~8 CFU/mL for 12 h.In the immersion challenge experiments,the composite adjuvant formulation combining chitosan quaternary ammonium salt particles and sodium alginate significantly increased the expression levels of immune-related cytokines,such as MHC-I,by 2 to 8 times,and improved the survival rate of turbot from 0%to 30%.Conclusion:The preparation process for chitosan and its quaternary ammonium salt particle adjuvants is suitable for scale-up production.Furthermore,the composite adjuvant of chitosan quaternary ammonium salt particles and sodium algin

关 键 词:大菱鲆 鳗弧菌 壳聚糖 口服疫苗 浸泡攻毒 

分 类 号:Q819[生物学—生物工程]

 

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