College of Veterinary Medicine (Institute of Comparative Medicine), Yangzhou University, 12 East Wenhui Road, Yangzhou, 225009, China.
Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Joint International Research Laboratory of Agriculture and Agri-Product Safety of Ministry of Education of China, Yangzhou, China.
BMC Vet Res. 2022 Aug 12;18(1):308. doi: 10.1186/s12917-022-03412-3.
Bacterial flagellin is a potent powerful adjuvant, which exerts its adjuvant activity by activating the Toll-like receptor 5 (TLR5) signaling pathway to induce host pro-inflammatory responses. Flagellin of Salmonella typhimurium (S. typhimurium) has shown strong adjuvant effects for a variety of vaccine candidates, however, the adjuvanticity of different serotypes of Escherichia coli (E. coli) flagellin (FliC) is unclear. To explore the adjuvant activity of different serotypes of E. coli flagellin, FliC, FliC, and FliC recombinant flagellins were prokaryotically-expressed and purified. The adjuvanticity of three recombinant flagellins was evaluated by analyzing their abilities to induce the IL-8 production in human colorectal adenocarcinoma (Caco-2) cells and the immune responses to co-administrated FaeG antigen in mice. Sequence analysis showed that the N-and C-terminal regions are highly conserved, whereas the central region is hypervariable. The TLR5 recognized site is identical among these three serotypes of flagellins. Coomassie blue staining SDS-PAGE showed the molecular mass of FliC, FliC, and FliC recombinant flagellin are 66 kDa, 64 kDa, and 68 kDa, which can be recognized by anti-FliC, FliC, and FliC serum, respectively. Moreover, the flagellin serotypes induced similar levels of IL-8 and TNF-α production in Caco-2 cells, anti-FaeG specific IgG antibodies in mice, and IL-4 production in mice spleen cells. Our results indicated that E. coli flagellins can be an adjuvant for vaccine candidates and that different serotypes of E. coli flagellins possess identical adjuvant effects.
细菌鞭毛蛋白是一种有效的佐剂，通过激活 Toll 样受体 5（TLR5）信号通路来诱导宿主炎症反应，发挥其佐剂活性。鼠伤寒沙门氏菌（S. typhimurium）的鞭毛蛋白已显示出对多种疫苗候选物的强烈佐剂作用，然而，不同血清型大肠杆菌（E. coli）鞭毛蛋白（FliC）的佐剂作用尚不清楚。为了探索不同血清型大肠杆菌鞭毛蛋白的佐剂活性，原核表达并纯化了 FliC、FliC 和 FliC 重组鞭毛蛋白。通过分析这三种重组鞭毛蛋白诱导人结直肠腺癌细胞（Caco-2）产生白细胞介素-8（IL-8）的能力以及与共同给药的 FaeG 抗原在小鼠中的免疫反应，评估了它们的佐剂活性。序列分析表明，N-和 C-末端区域高度保守，而中心区域高度可变。这三种鞭毛蛋白的 TLR5 识别位点相同。考马斯亮蓝染色 SDS-PAGE 显示 FliC、FliC 和 FliC 重组鞭毛蛋白的分子量分别为 66 kDa、64 kDa 和 68 kDa，可分别被抗 FliC、FliC 和 FliC 血清识别。此外，鞭毛蛋白血清型在 Caco-2 细胞中诱导产生相似水平的白细胞介素-8 和肿瘤坏死因子-α、小鼠抗 FaeG 特异性 IgG 抗体和小鼠脾细胞白细胞介素-4。我们的结果表明，大肠杆菌鞭毛蛋白可作为疫苗候选物的佐剂，不同血清型的大肠杆菌鞭毛蛋白具有相同的佐剂作用。
