Preparation of Ferritin Nanoparticles Displaying the Dominant Neutralizing Epitopes of Foot-and-mouth disease virus and Their Immunogenicity Evaluation in Mouse (Mus musculus) Models
LU Bing-Zhou1,2, RU Yi2, HAO Rong-Zeng2, LI Ya-Jun2, YANG Yang2, YANG Rui1, MAO Yu-Han2, JIANG Cheng-Hui2, TAO Shi-Yu1, ZHENG Hai-Xue2, YU Si-Jiu1, CUI Yan1,*
1 College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China; 2 Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences/State Key Laboratory of Veterinary Etiological Biology, Lanzhou 730046, China
Abstract：Nanoparticles (NPs) can display multiple copies of antigen on the surface and simulate the structural morphology of viruses, which is more conducive to uptake by immune cells and promote antigen presentation, thus enhancing protective efficacy. This study aimed to generate ferritin nanoparticles harboring a dominant neutralizing epitope of Foot-and-mouth disease virus (FMDV) by using the Escherichia coli expression system, and to evaluate its immunogenicity. The recombinant expression plasmid pET28a-FBT was constructed based on the Helicobacter pylori ferritin, the dominant neutralizing epitope sequence of FMDV was inserted between the 33th and 34th amino acids of ferritin, and the T cell epitope sequence was inserted at the N-terminus of ferritin. The recombinant expression plasmid pET28a-Fer expressing Fer protein was used as control. The recombinant plasmid pET28a-Fer and pET28a-FBT were transformed into Escherichia coli competent cells, the target proteins were obtained by induced expression and size exclusion chromatography purification, and identified using SDS-PAGE, Western blot, particle size analysis, and transmission electron microscopy, then evaluated immunogenicity by immunizing BALB/c mice (Mus muculus). The results showed that the recombinant protein Fer and FBT were successfully prepared by using prokaryotic expression system. The recombinant protein FBT could be recognized specifically by FMDV positive serum. After 14 d of immunization with FBT, serum antibodies were produced and continued to rise, and the neutralizing antibody titer could reach up to 1∶128 on 35 d. At the same time, the proliferation level of spleen lymphocytes and the secretion level of interleukin 2 (IL-2), IL-4, IL-10, and interferon-γ (IFN-γ) in FBT immunized mice were significantly higher than the control (P<0.05). This study prepared ferritin nanoparticles displaying FMDV neutralizing epitopes with good immunogenicity, providing technical support for the development of a novel foot-and-mouth disease engineered vaccine.
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