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| 免疫增强剂CVC1302诱导高亲和力抗体机制分析 |
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杜露平1,2,3*,鲁海燕1,2,3,侯立婷2,3,于晓明2,3,程海卫2,3,张元鹏2,3,陈瑾1,2,3,郑其升1,2,3,甘芳1
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| (1.南京农业大学 动物医学院, 南京 210095;2.江苏省农业科学院 动物免疫工程研究所/国家兽用生物制品工程中心/江苏省食品质量安全重点实验室, 南京 210014;3.兽用生物制品(泰州)国泰技术创新中心, 江苏 泰州 225300) |
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| 摘要: |
| 为探究免疫增强剂CVC1302诱导机体产生高亲和力抗体的免疫机制，本研究利用4-羟基-3-硝基苯乙酰基耦联鸡卵白蛋白(NP-OVA)与免疫增强剂CVC1302配伍后，与ISA206佐剂乳化制备疫苗。将33只6周龄BALB/c雌性小鼠随机分为3组，分别后腿肌肉注射NP、NP-ISA206和NP-CVC1302-ISA206，每只100 μL疫苗，50 μg NP-OVA/只，利用ELISA检测高亲和力NP特异性抗体水平；利用流式细胞仪分析生发中心B细胞增殖水平、生发中心 B细胞诱导活化的胞苷脱氨酶(AID)表达水平；利用荧光定量PCR检测AID基因和HoxC4基因转录水平。结果表明:1)免疫增强剂CVC1302可诱导小鼠产生高亲和力的NP特异性抗体。2)免疫增强剂CVC1302可显著提升抗原特异性生发中心 B细胞数量占总B细胞数量比例。3)免疫增强剂CVC1302可显著增加生发中心 B细胞HoxC4基因转录水平。4)免疫增强剂CVC1302可显著增强生发中心B细胞AID蛋白表达水平。生发中心B细胞表达的AID可针对B细胞免疫球蛋白序列可变区进行点突变，发生体细胞高频突变，提升B细胞BCR针对抗原的亲和力，进而在辅助性T细胞的阳性选择下分化为长寿浆细胞，持续性分泌高亲和力抗体，因此本研究推断免疫增强剂CVC1302依赖于正向调控AID诱导机体产生高亲和力抗体，为机体提供免疫保护力，以抵抗病原体的感染，为后续新型免疫增强剂的研制提供思路和理论基础。 |
| 关键词: 免疫增强剂CVC1302 高亲和力抗体 AID 生发中心B细胞 HoxC4 |
| DOI：10.11841/j.issn.1007-4333.2023.10.14 |
| 投稿时间：2022-12-07 |
| 基金项目:十四五重点研发专项(2022YFD1800800)；国家自然科学基金项目(32102690)；江苏省农业自主创新专项(CX(21)3135) |
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| Analysis of the mechanism of immunopotentiator CVC1302 inducing high affinity antibody |
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DU Luping1,2,3*,LU Haiyan1,2,3,HOU Liting2,3,YU Xiaoming2,3,CHENG Haiwei2,3,ZHANG Yuanpeng2,3,CHEN Jin1,2,3,ZHENG Qisheng1,2,3,GAN Fang1
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| (1.College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China;2.Institute of Veterinary Immunology & Engineering, Jiangsu Academy of Agricultural Sciences/National Research Center ofEngineering and Technology for Veterinary Biologicals/Jiangsu Key Laboratory for Food Quality and Safety-State KeyLaboratory Cultivation Base, Ministry of Science and Technology, Nanjing 210014, China;3.GuoTai(Taizhou)Center of Technology Innovation for Veterinary Biologicals, Taizhou 210014, China) |
| Abstract: |
| In order to elucidate the immune-mechanism of immunopotentiator CVC1302 inducing high affinity antibody, NP-OVA, as model antigen, mixed with CVC1302, then emulsified with ISA206, was prepared for vaccine. 33 Six-week-old BALB/c female mice were divided into 3 groups and immunized with NP-CVC1302-ISA206, NP-ISA206 or NP, each mouse immunized with 100 μL vaccine, as well as 50 μg NP-OVA. The levels of high-affinity NP-specific antibody were detected by ELISA, the percentages of NP-specific germinal center(GC)B cells in B cells and expression levels of AID in GC B cells were analyzed by flow cytometry. GC B cells were harvested by FACS sorting. The transcriptional levels of AID and HoxC4 in GC B cells were analyzed by real-time PCR. The results showed that: 1)Immunopotentiator CVC1302 induced high affinity NP-specific antibodies. 2)CVC1302 induced higher percentage of NP-specific GC B cells in B cells. 3)CVC1302 improved transcriptional levels of HoxC4 in GC B cells. 4)CVC1302 enhanced the expression levels of AID in GC B cells. Considering AID in GC B cells inducing high affinity antibodies through point mutation, then GC B cells differentiated into long-term plasma cells under the positive selection of follicular helper T cells, we concluded that CVC1302 induced high affinity antibody dependent on positive control of AID, which shed light on the development of new immunopotentiators. |
| Key words: immunopotentiator CVC1302 high affinity antibody AID GC B cell HoxC4 |
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