The fundamental study for polymyxins as novel and safe mucosal adjuvants

目前迫切需要开发安全有效的佐剂来增强疫苗诱导的抗原特异性免疫反应。在这里，我们证明了临床使用的多肽抗生素多粘菌素B(PMB)和粘菌素(CL)与卵清蛋白(OVA)一起鼻腔免疫，以剂量依赖的方式增加了OVA特异性的体液免疫反应，无论是在粘膜还是在全身。在8个月的观察中，通过加强免疫而增强的免疫力持续存在。此外，与单独使用OVA的小鼠相比，OVA+不同剂量的PMB或CL鼻腔免疫的小鼠在鼻腔和嗅球中既没有炎症反应，也没有肾脏损害。这些数据表明，多粘菌素可能是一种新的、安全的粘膜佐剂来诱导体液免疫反应。多粘菌素的佐剂性不依赖于其杀菌活性所释放的内毒素，在脂多糖(LPS)反应迟钝和敏感的小鼠中，PMB的增强作用相似。然而，尽管存在预先存在的抗PMB抗体，我们观察到多粘菌素在鼻腔给药时并没有减少其佐剂功能。此外，OVA+PMB或CL鼻腔免疫的小鼠的OVA特异性抗体效价显著高于多粘菌素类似物，如多粘菌素B、多粘菌素甲烷磺酸盐。PMB或CL刺激的肥大细胞培养上清液中释放的β-氨基己糖苷酶和组胺水平也明显高于其类似物刺激的水平。这些结果表明，疏水碳链和亲水性阳离子环肽都参与了PMB和CL的粘膜佐剂作用

There is currently an urgent need to develop safe and effective adjuvants for enhancing vaccine-induced antigen -specific immune responses. We demonstrate here that intranasal immunization with clinically used polypeptide antibiotics, polymyxin B (PMB) and colistin (CL), along with ovalbumin (OVA), increases OVA-specific humoral immune responses in a dose-depe ndently manner at both mucosal and systemic compartments. Enhanced immunity by boosting was found to persist during 8 months of observation. Moreover, mice intranasally immunized with OVA plus various doses of PMB or CL showed neither inflammatory responses in the nasal cavity and olfactory bulbs nor renal damages, compared to those given OVA alone. These data suggest that polymyxins may serve as novel and safe mucosal adjuvants to induce humoral immune responses. The polymyxin adjuvanticity was found to be independent of endotoxins liberated by its bactericidal activity, as indicated by similar enhancing effects of PMB in lipopolysaccharide (LPS)-hyporesponsive and LPS -susceptible mice. However, despite the presence of preexisting anti-PMB antibodies, we ob served no reduction in the adjuvant function of polymyxins when they were given intranasally. Furthermore, the titers of OVA-specific Abs in mice intranasally immunized with OVA plus PMB or CL were significantly higher than those in mice administered with polymyxin analogues, such as polymyxin B nonapeptide and colistin methanesulfonate. The levels of released β-hexosaminidase and histamine in mast cell culture supernatants stimulated by PMB or CL were also significantly higher than those stimulated by their analogues. These results suggest that both the hydrophobic carbon chain and hydrophilic cationic cyclic peptide contribute to the mucosal adjuvanticity of PMB and CL