Department of Biotechnology, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria, INIA, Madrid, Spain.
PLoS One. 2012;7(11):e48466. doi: 10.1371/journal.pone.0048466. Epub 2012 Nov 14.
Flagellins evoke strong innate and adaptive immune responses. These proteins may play a key role as radioprotectors, exert antitumoral activity in certain types of tumor and reduce graft-versus-host disease in allogeneic hematopoietic stem cell transplant recipients. Notwithstanding, flagellins are highly immunogenic, and repeated use leads to their neutralization by systemic antibodies. This neutralization is not prevented by using functional deleted flagellins. These observations led us to explore the possibility of preventing initial neutralization by means of another functional flagellin that does not belong to common pathogenic bacteria but that has the capacity to activate TLR5. Here we characterized the functional capacity of the two-phase Marinobacter algicola (MA)-derived flagellins (F and FR) as systemic and mucosal adjuvants and compared their performance with that of Salmonella typhimurium (STF) flagellins (FljB and FliC). We also report for the first time on the in vitro and in vivo capacity of various flagellins to trigger TLR5 activation in the presence of species-specific anti-flagellin antibodies, the cross-neutralization mediated by these antibodies, and the sequential use of these flagellins for TLR5 activation. Our results showed that MA flagellins behave in a similar way to STF ones, inducing pro-inflammatory cytokines (IL8, CCL20, CCL2) and evoking a strong in vivo antibody response against a model epitope. More importantly, MA flagellins were fully functional, in vitro or in vivo, in the presence of a high concentration of neutralizing anti-flagellin STF antibodies, and STF flagellin was not inhibited by neutralizing anti-flagellin MA antibodies. The use of active flagellins from distinct bacteria could be a useful approach to prevent systemic neutralization of this group of adjuvants and to facilitate the rational design of flagellin-based vaccines and/or other therapeutic treatments (against ischemia, acute renal failure, tumors, ionizing radiations and also to improve the outcome of bone marrow transplants).
鞭毛蛋白会引起强烈的先天和适应性免疫反应。这些蛋白质可能在作为辐射防护剂方面发挥关键作用，在某些类型的肿瘤中发挥抗肿瘤活性，并减少异基因造血干细胞移植受者中的移植物抗宿主病。尽管如此，鞭毛蛋白具有高度的免疫原性，重复使用会导致全身性抗体中和它们。使用功能缺失的鞭毛蛋白并不能防止这种中和。这些观察结果促使我们探索另一种不属于常见病原菌但具有激活 TLR5 能力的功能性鞭毛蛋白来预防初始中和的可能性。在这里，我们描述了 Marinobacter algicola（MA）衍生的鞭毛蛋白（F 和 FR）作为全身性和黏膜佐剂的功能能力，并将其性能与 Salmonella typhimurium（STF）鞭毛蛋白（FljB 和 FliC）进行了比较。我们还首次报道了各种鞭毛蛋白在存在特异性抗鞭毛蛋白抗体的情况下触发 TLR5 激活的体外和体内能力、这些抗体介导的交叉中和以及这些鞭毛蛋白用于 TLR5 激活的顺序使用。我们的研究结果表明，MA 鞭毛蛋白的行为与 STF 鞭毛蛋白相似，可诱导促炎细胞因子（IL8、CCL20、CCL2）并引发针对模型表位的强烈体内抗体反应。更重要的是，MA 鞭毛蛋白在存在高浓度中和性抗鞭毛蛋白 STF 抗体的情况下，在体外或体内均具有完全功能，而 STF 鞭毛蛋白不受中和性抗鞭毛蛋白 MA 抗体的抑制。使用来自不同细菌的活性鞭毛蛋白可能是一种有用的方法，可以防止此类佐剂的全身性中和，并有助于基于鞭毛蛋白的疫苗和/或其他治疗方法（针对缺血、急性肾衰竭、肿瘤、电离辐射以及改善骨髓移植的结果）的合理设计。
