College of Life Sciences, China Jiliang University, Zhejiang, 310018, Hangzhou, China.
Institute of Animal Husbandry and Veterinary Science, Zhejiang Academy of Agricultural Sciences, Zhejiang, 310021, Hangzhou, China.
Appl Microbiol Biotechnol. 2024 Feb 26;108(1):238. doi: 10.1007/s00253-024-13060-2.
Pasteurella multocida is an important bacterial pathogen that can cause diseases in both animals and humans. Its elevated morbidity and mortality rates in animals result in substantial economic repercussions within the livestock industry. The prevention of diseases caused by P. multocida through immunization is impeded by the absence of a safe and effective vaccine. Outer membrane vesicles (OMVs) secreted from the outer membrane of Gram-negative bacteria are spherical vesicular structures that encompass an array of periplasmic components in conjunction with a diverse assortment of lipids and proteins. These vesicles can induce antibacterial immune responses within the host. P. multocida has been shown to produce OMVs. Nonetheless, the precise characteristics and immunomodulatory functions of P. multocida OMVs have not been fully elucidated. In this study, OMVs were isolated from P. multocida using an ultrafiltration concentration technique, and their morphology, protein constitution, and immunomodulatory properties in RAW264.7 cells were studied. Transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA) revealed that the OMVs exhibited typical spherical and bilayered lipid vesicular architecture, exhibiting an average diameter of approximately 147.5 nm. The yield of OMVs was 2.6 × 10 particles/mL. Proteomic analysis revealed a high abundance of membrane-associated proteins within P. multocida OMVs, with the capability to instigate the host's immune response. Furthermore, OMVs stimulated the proliferation and cellular uptake of macrophages and triggered the secretion of cytokines, such as TNF-ɑ, IL-1β, IL-6, IL-10, and TGF-β1. Consequently, our results indicated that OMVs from P. multocida could directly interact with macrophages and regulate their immune function in vitro. These results supported the prospective applicability of P. multocida OMVs as a platform in the context of vaccine development. KEY POINTS: • Preparation and characterization of P. multocida OMVs. • P. multocida OMVs possess a range of antigens and lipoproteins associated with the activation of the immune system. • P. multocida OMVs can activate the proliferation, internalization, and cytokine secretion of macrophages in vitro.
多杀巴斯德菌是一种重要的细菌性病原体，可导致动物和人类患病。其在动物中的高发病率和死亡率给畜牧业带来了巨大的经济影响。由于缺乏安全有效的疫苗，通过免疫预防多杀巴斯德菌引起的疾病受到阻碍。革兰氏阴性菌外膜囊泡（OMVs）是从外膜分泌的球形囊泡结构，包含一系列周质成分以及多种脂质和蛋白质。这些囊泡可以在宿主中诱导抗菌免疫反应。多杀巴斯德菌已被证明可以产生 OMVs。然而，多杀巴斯德菌 OMVs 的精确特征和免疫调节功能尚未完全阐明。在这项研究中，使用超滤浓缩技术从多杀巴斯德菌中分离出 OMVs，并研究了它们在 RAW264.7 细胞中的形态、蛋白质组成和免疫调节特性。透射电子显微镜（TEM）和纳米颗粒跟踪分析（NTA）显示，OMVs 呈现出典型的球形双层脂质囊泡结构，平均直径约为 147.5nm。OMVs 的产量为 2.6×10 个/mL。蛋白质组学分析显示，多杀巴斯德菌 OMVs 中富含膜相关蛋白，具有引发宿主免疫反应的能力。此外，OMVs 刺激巨噬细胞的增殖和细胞摄取，并触发细胞因子如 TNF-ɑ、IL-1β、IL-6、IL-10 和 TGF-β1 的分泌。因此，我们的结果表明，多杀巴斯德菌的 OMVs 可以直接与巨噬细胞相互作用，并在体外调节其免疫功能。这些结果支持多杀巴斯德菌 OMVs 作为疫苗开发平台的潜在应用。 要点： • 多杀巴斯德菌 OMVs 的制备和表征。 • 多杀巴斯德菌 OMVs 具有一系列与免疫系统激活相关的抗原和脂蛋白。 • 多杀巴斯德菌 OMVs 可以在体外激活巨噬细胞的增殖、内化和细胞因子分泌。
