The role of LPS-TLR4 signaling in live vaccine-induced protective responses

活疫苗在提供有效的预防后续疫苗方面有很长的历史 具有感染力的挑战。然而，在许多情况下，导致保护的机制是 定义不明确。我们的目标是定义革兰氏阴性的免疫保护机制 以新型鼠疫耶尔森氏菌为模型的细菌疫苗株 系统。革兰氏阴性菌鼠疫耶尔森氏菌是鼠疫的病原体。 目前还没有获得许可的鼠疫疫苗，而探索性疫苗已经 预防肺炎的能力不同，这是生物恐怖后的预期形式 进攻。我们开发了一种新的方法来产生有效的疫苗株 用于预防鼠疫和潜在的其他微生物，基于 增强固有细菌Toll样受体-4介导的佐剂活性。 与其他各种革兰氏阴性细菌相似，鼠疫杆菌产生一种脂多糖。 (LPs)在37℃时刺激能力低。TLR4是细胞内的内毒素受体，通过其 脂类A。我们培育了一株表达LpxL的新的鼠疫耶尔森氏菌。 生物合成酶，并发现这能在37℃产生有效的内毒素。 由于先天抗菌素的诱导，通过外周接种使小鼠无毒 通过TLR4的免疫机制，这一途径也与强佐剂作用相关。 我们的结果表明，用鼠疫杆菌LpxL株免疫小鼠可诱导充分 对强毒小鼠皮下和鼻腔攻击的保护作用 细菌，模仿腺鼠疫和肺鼠疫。我们的主要假设是 许多含有增强效力的内毒素的活细菌疫苗菌株是 有效的疫苗，增加的TLR4信号将提供增强的 适应性免疫反应。我们建议确定影响生物多样性的机制 使用产生有效内毒素的活和灭活鼠疫杆菌的疫苗效力，通过与 没有增加TLR4刺激的菌株，测试体外和体内反应。 现有的和新的减毒株都将使用。依靠一次枝晶 细胞和基因缺陷小鼠，我们将研究TLR信号通路导致 树突状细胞在体内和体外的激活，抗原提呈和T细胞激活。我们 将分析疫苗对皮下和鼻腔感染的效果。这个 这些研究的完成将提供有关疫苗接种机制的信息 革兰氏阴性菌感染的菌株可能会起作用。掺入TLR-刺激物 直接免疫逃避病原体的佐剂活性可能构成一种新的方法 用于减毒和疫苗的生成。

Live vaccines have a long history for providing efficient protection against subsequent infectious challenge. However, the mechanisms leading to protection are in many cases not well defined. Our goal is to define mechanisms for immune protection by Gramnegative bacterial vaccine strains, using novel Yersinia pestis strains as model systems. The gram-negative bacterium Yersinia pestis is the causative agent of plague. Currently there is no available licensed plague vaccine, and exploratory vaccines have variable ability to protect against pneumonic disease, the form expected after a bioterror attack. We have developed a new method for the generation of efficient vaccine strains for protection against plague and potentially other microorganisms, based upon enhancement of inherent bacterial Toll-like receptor (TLR)-4 mediated adjuvant activity. Similar to various other gram-negative bacteria, Y. pestis produces a lipopolysaccharide (LPS) with low stimulatory ability at 37¿C. TLR4 is the cellular receptor for LPS via its lipid A. We generated a new Y. pestis strain expressing LpxL, an E. coli lipid A biosynthesis enzyme, and found this to produce a potent LPS at 37¿C. This strain is avirulent in mice by peripheral inoculation, due to induction of antibacterial innate immune mechanisms via TLR4, a pathway also associated with strong adjuvant effects. Our results indicate that vaccination of mice with the Y. pestis LpxL strain induces full protection against both subcutaneous and intranasal challenge of mice with virulent bacteria, mimicking bubonic and pneumonic plague. Our main hypotheses are that many live bacterial vaccine strains containing LPS with increased potency are efficient vaccines, and that the increased TLR4 signaling will provide enhanced adaptive immune responses. We propose to determine mechanisms influencing the vaccine efficacy using live and killed Y. pestis producing a potent LPS, by comparing to strains without increased TLR4 stimulation, testing both in vitro and in vivo responses. Both existing and novel attenuated strains will be used. Relying on primary dendritic cells and genetically deficent mice, we will study TLR signaling pathways leading to dendritic cell activation in vivo and in vitro, antigen presentation and T cell activation. We will analyze vaccine effects against both subcutaneous and intranasal infection. The completion of these studies will provide information on the mechanism by which vaccine strains towards Gram-negative infections may act. Incorporation of TLR-stimulating adjuvant activity directly into immune-evading pathogens may constitute a novel method for attenuation and generation of vaccines.