Department of Microbiology, Faculty of Science, Mahidol University, Bangkok, Thailand.
Department of Microbiology and Immunology, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand.
PLoS One. 2018 Jun 14;13(6):e0198906. doi: 10.1371/journal.pone.0198906. eCollection 2018.
Burkholderia pseudomallei is a flagellated Gram-negative bacterium which is the causative agent of melioidosis. The disease poses a major public health problem in tropical regions and diabetes is a major risk factor. The high mortality rate of melioidosis is associated with severe sepsis which involves the overwhelming production of pro-inflammatory cytokines. Bacterial flagellar protein (flagellin) activates Toll-like receptor 5 (TLR5)-mediated innate immune signaling pathways and induces adaptive immune response. However, previous studies of TLR5 signaling in melioidosis have been performed using recombinant flagellin from Salmonella Typhimurium instead of B. pseudomallei. This study aimed to investigate human innate immune response and antibody response against a recombinant B. pseudomallei flagellin (rFliC). We prepared B. pseudomallei rFliC and used it to stimulate HEK-BlueTM-hTLR5 and THP1-DualTM cells to assess TLR5 activation. Subsequently, whole blood stimulation assays with rFliC were performed ex vivo. TLR5-flagellin interactions trigger activation of transcription factor NF-κB in HEK-BlueTM-hTLR5 cells. Pro-inflammatory cytokine (IL-1β, IL-6, and TNF-α) productions from whole blood in response to rFliC differed between fourteen healthy individuals. The levels of these cytokines changed in a dose and time-dependent manner. ELISA was used to determine rFliC-specific antibodies in serum samples from different groups of melioidosis patients and healthy subjects. IgG antibody to rFliC in melioidosis patients with diabetes were higher compared with non-diabetic patients. Our results show that B. pseudomallei flagellin is a potent immune stimulator and that the immune responses to rFliC are different among individuals. This may provide valuable insights toward the potential use of rFliC in vaccine development.
类鼻疽伯克霍尔德菌是一种具有鞭毛的革兰氏阴性细菌，是类鼻疽病的病原体。该疾病在热带地区对公共卫生构成重大威胁，糖尿病是一个主要的危险因素。类鼻疽病的高死亡率与严重败血症有关，严重败血症涉及过度产生促炎细胞因子。细菌鞭毛蛋白（鞭毛蛋白）激活 Toll 样受体 5（TLR5）介导的先天免疫信号通路，并诱导适应性免疫反应。然而，之前对类鼻疽病中 TLR5 信号的研究使用的是重组沙门氏菌鞭毛蛋白（鞭毛蛋白）而不是类鼻疽伯克霍尔德菌。本研究旨在研究人类对重组类鼻疽伯克霍尔德菌鞭毛蛋白（rFliC）的固有免疫反应和抗体反应。我们制备了类鼻疽伯克霍尔德菌 rFliC，并使用它来刺激 HEK-BlueTM-hTLR5 和 THP1-DualTM 细胞，以评估 TLR5 的激活。随后，在体外进行 rFliC 的全血刺激试验。TLR5-鞭毛蛋白相互作用触发转录因子 NF-κB 在 HEK-BlueTM-hTLR5 细胞中的激活。rFliC 对全血中促炎细胞因子（IL-1β、IL-6 和 TNF-α）的刺激在 14 名健康个体之间存在差异。这些细胞因子的水平呈剂量和时间依赖性变化。ELISA 用于测定不同组类鼻疽病患者和健康受试者血清样本中的 rFliC 特异性抗体。糖尿病类鼻疽病患者的 rFliC 特异性 IgG 抗体水平高于非糖尿病患者。我们的结果表明，类鼻疽伯克霍尔德菌鞭毛蛋白是一种有效的免疫刺激剂，并且个体对 rFliC 的免疫反应不同。这可能为 rFliC 在疫苗开发中的潜在用途提供有价值的见解。
