Vaccine strategies that target cytolic CD4 T cells to the lung.

描述（由申请人提供）：呼吸道病毒感染仍然是引起发病率和死亡率的主要健康问题，特别是在老年人和免疫功能低下的个体中。目前的流感疫苗依赖于诱导中和抗体来包裹蛋白质；然而，这些疫苗对血清学上不同的病毒株和具有大流行潜力的病毒株无效。因此，开发针对T细胞免疫的流感疫苗是极其重要的，然而，指导T细胞运输到肺部的机制以及与保护相关的T细胞效应机制尚未完全确定。此外，目前尚不清楚哪种佐剂或疫苗制剂最适合于持续记忆的初始有效T细胞反应。为了解决这些知识上的空白，我们将使用流感感染小鼠模型和特异性流感H1N1的CD4 T细胞受体（TCR）转基因（tg）小鼠来追踪CD4 T细胞效应物的发展，这些效应物运输到肺部，持续记忆并有助于接种疫苗后的保护。我们之前的研究和初步数据表明，CD4 T细胞在流感感染后获得穿孔素介导的细胞毒性，并以抗体独立的方式保护细胞免受致命攻击。本建议的目的是确定哪种疫苗策略可引起细胞溶解性CD4 T细胞迁移到肺部并对致命流感感染提供保护。我们将重点关注toll样受体（TLR）配体作为佐剂，因为它们具有动员树突状细胞（DC）的能力，树突状细胞是T细胞反应的有效激活剂。在Aim 1中，我们将确定TLR配体在体外和体内促进II类限制性细胞溶解活性中的作用。第一种方法是从肺中分离DC细胞群，用TLR配体激活，与TCR Tg CD4细胞共培养，并确定是否在体外产生溶细胞CD4细胞。第二种方法将是通过将TCR Tg CD4细胞转移到小鼠体内，经呼吸道接种并分析肺部CD4反应来证实TLR激动剂在体内的辅助作用。在目的2中，我们将确定疫苗诱导的细胞溶解CD4细胞是否分化到记忆阶段，并有助于预防致命的流感感染。我们将使用TLR配体诱导细胞溶解CD4细胞作为相关的保护，并测试这些佐剂是否诱导记忆群体并提供对致命流感挑战的保护。最后，使用穿孔素缺陷的CD4 T细胞和B细胞缺陷小鼠，我们将测试穿孔素依赖的细胞溶解活性是否需要由CD4 T细胞介导的抗体独立保护。

DESCRIPTION (provided by applicant): Respiratory viral infections remain a major health concern causing morbidity and mortality, especially in elderly and immunocompromised individuals. Current vaccines against influenza rely on inducing neutralizing antibodies to coat proteins; however, these vaccines are ineffective against serologically distinct viral strains and strains that have pandemic potential. Therefore, it is of extreme importance to develop influenza vaccines that target T cell immunity, however, the mechanisms that direct T cell trafficking to the lung and the T cell effector mechanisms that correlate with protection have not been fully defined. In addition, it is not known which adjuvants or vaccine preparations are best suited to prime effective T cell responses that persist to memory. To address these gaps in our knowledge, we will use a mouse model of influenza infection and CD4 T cell receptor (TCR) transgenic (tg) mice specific for influenza H1N1 to track the development of CD4 T cell effectors that traffic to the lung, persist to memory and contribute to protection following vaccination. Our previous studies and preliminary data demonstrate that CD4 T cells acquire perforin mediated cytotoxicity following influenza infection and protect against lethal challenge in an antibody independent manner. The objective of this proposal is to determine which vaccine strategies elicit cytolytic CD4 T cells that migrate to the lung and afford protection to lethal influenza infection. We will focus on toll like receptor (TLR) ligands as adjuvants because of their ability to mobilize dendritic cells (DC), potent activators of T cell responses. In Aim 1, we will determine the role of TLR ligands in promoting class II restricted cytolytic activity in vitro and in vivo. The first approach will be to isolate DC populations from the lung, activate with TLR ligands, co-culture with TCR Tg CD4 cells and determine if cytolytic CD4 cells are generated in vitro. The second approach will be to confirm the adjuvant effect of TLR agonists in vivo by transferring naive TCR Tg CD4 cells into mice, vaccinating via the respiratory tract and analyzing the CD4 response in the lung. In Aim 2 we will determine whether vaccine induced cytolytic CD4 cells differentiate to the memory stage and contribute to protection against lethal influenza infection. We will use the TLR ligands that elicit cytolytic CD4 cells as a correlate of protection and test whether those adjuvants induce a memory population and provide protection against a lethal influenza challenge. Finally, using perforin deficient CD4 T cells and B cell deficient mice, we will test whether perforin dependent cytolytic activity is required for antibody independent protection mediated by CD4 T cells. PUBLIC HEALTH RELEVANCE: Respiratory infections such as influenza pose a major public health threat. The proposed studies will further our understanding of how CD4 T cell immunity develops and persists in the lung in response to vaccination with adjuvant preparations that activate the innate immune response. This information will be important for developing novel vaccine strategies that target cell mediated immunity and provide more universal protection against emerging, serologically distinct influenza strains such as H5N1 and H1N1.