Zhang Yue, Ru Yi, Hao Rongzeng, Yang Yang, Zhao Longhe, Li Yajun, Yang Rui, Lu Bingzhou, Zheng Haixue
State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou University, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730000, Gansu, China.
College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, Gansu, China.
Sheng Wu Gong Cheng Xue Bao. 2024 Dec 25;40(12):4509-4520. doi: 10.13345/j.cjb.240354.
This study developed ferritin-based nanoparticles carrying the African swine fever virus (ASFV) p30 protein and evaluated their immunogenicity, aiming to provide an experimental basis for the research on nanoparticle vaccines against ASFV. Initially, the gene sequences encoding the p30 protein and SpyTag were fused and inserted into the pCold-I vector to create the pCold-p30 plasmid. The gene sequences encoding SpyCatcher and ferritin were fused and then inserted into the pET-28a(+) vector to produce the pET-F-np plasmid. Both plasmids were expressed in upon induction. Subsequently, the affinity chromatography-purified p30 protein was conjugated with ferritin , and the p30-ferritin (F-p30) nanoparticles were purified by size-exclusion chromatography. The morphology and structural integrity of F-p30 nanoparticles were examined by a particle size analyzer and transmission electron microscopy. Mice were immunized with F-p30 nanoparticles, and the humoral and cellular immune responses were assessed. The results showed that F-p30 nanoparticles were successfully prepared, with the particle size of approximately 20 nm. F-p30 nanoparticles were efficiently internalized by bone marrow-derived dendritic cells (BMDCs) cells . Compared with the p30 protein alone, F-p30 nanoparticles induced elevated levels of specific antibodies and cytokines in mice and stimulated the proliferation of follicular helper T cell (T) and germinal center B cell (GCB) in lymph nodes as well as CD4 and CD8 T cells in the spleen. In conclusion, we successfully prepared F-p30 nanoparticles which significantly enhanced the immunogenicity of p30 protein, giving insights into the development of vaccines against ASFV.
本研究开发了携带非洲猪瘟病毒（ASFV）p30蛋白的铁蛋白基纳米颗粒，并评估了它们的免疫原性，旨在为抗ASFV纳米颗粒疫苗的研究提供实验依据。最初，将编码p30蛋白和SpyTag的基因序列融合并插入pCold-I载体中，构建pCold-p30质粒。将编码SpyCatcher和铁蛋白的基因序列融合，然后插入pET-28a(+)载体中，构建pET-F-np质粒。两种质粒经诱导后均在[具体表达宿主未给出]中表达。随后，通过亲和层析纯化的p30蛋白与铁蛋白[此处未提及具体结合方式或其他相关信息]结合，并通过尺寸排阻色谱法纯化p30-铁蛋白（F-p30）纳米颗粒。通过粒度分析仪和透射电子显微镜检查F-p30纳米颗粒的形态和结构完整性。用F-p30纳米颗粒免疫小鼠，并评估其体液免疫和细胞免疫反应。结果表明，成功制备了粒径约为20 nm的F-p30纳米颗粒。F-p30纳米颗粒能被骨髓来源的树突状细胞（BMDCs）有效内化。与单独的p30蛋白相比，F-p30纳米颗粒在小鼠体内诱导产生了更高水平的特异性抗体和细胞因子，并刺激了淋巴结中滤泡辅助性T细胞（Tfh）和生发中心B细胞（GCB）以及脾脏中CD4和CD8 T细胞的增殖。总之，我们成功制备了F-p30纳米颗粒，其显著增强了p30蛋白的免疫原性，为抗ASFV疫苗的开发提供了思路。
