Priming of Antifungal T-Cells at Mucosal and Systemic Sites

描述（由申请人提供）：真菌病原体和大多数传染性病原体从粘膜表面进入人体。粘膜免疫启动诱导全身和粘膜抵抗，被认为是预防或治疗由粘膜表面引起的感染的理想策略。系统启动，反过来，通常诱导较差的粘膜免疫，被认为是不理想的粘膜感染。与此相反，我们发现皮炎芽孢菌减毒活疫苗株可以保护小鼠免受致命肺部感染（通过Th1 CD4+ T细胞），当减毒株仅通过皮下途径而不是通过呼吸途径传递时。在这里，我们将研究这两种抗原（Ag）递送途径在启动和召回阶段触发CD4+ T细胞的基本机制。我们将努力揭示导致T细胞在皮肤和肺中启动有效或失败的细胞和分子机制。T细胞受体（TCR）转基因（Tg）系统可以说是破解体内克隆T细胞激活的最佳免疫学工具，但目前还没有这样的试剂可用于研究T细胞对二态真菌的免疫。因此，我们建议创建一个胚菌特异性CD4+ TCR Tg小鼠来监测和分析ag特异性CD4+ T细胞在这两种免疫不同环境下的活化。我们假设Ag的皮下递送会激活皮肤引流淋巴结中的TCR Tg，胚细胞特异性CD4+ t细胞成为活化的Th1效应细胞，并在回忆时保护肺部免受攻击。另一方面，粘膜启动将导致产生功能受损的Th1效应器，无法提供抵抗挑战的能力。我们的具体目标是：1)开发一种过继性转移系统，在体内监测囊胚特异性CD4+ T细胞。真菌特异性TCR Tg小鼠将从T细胞克隆中创建，该T细胞克隆可保护小鼠免受实验感染。TCR Tg细胞将用于建立一个自体过继转移系统，以跟踪皮肤和肺中启动阶段保护性ag特异性T细胞的表型和功能特性。2)确定粘膜启动是否以及如何不能产生ag特异性Th1 CD4+细胞。TCR Tg细胞将让我们追踪Ag暴露于呼吸道粘膜后CD4+ T细胞反应的关键阶段：在启动和回忆阶段，肺和引流淋巴结的募集、激活、增殖和分化。这些事件将与皮肤中银暴露后的事件进行比较/对比。

DESCRIPTION (provided by applicant): Fungal pathogens and most infectious agents enter the body at mucosal surfaces. Mucosal immune priming induces both systemic and mucosal resistance, and is believed to be the ideal strategy to prevent or treat infections that arise from a mucosal surface. Systemic priming, in turn, usually induces poor mucosal immunity and is judged to be less desirable against mucosal infections. Contrary to this paradigm, we find that a live-attenuated, vaccine strain of Blastomyces dermatitidis can protect mice against a lethal pulmonary infection (via Th1 CD4+ T cells) when the attenuated strain is delivered only by the subcutaneous route, and not by the respiratory route. Here, we will investigate the fundamental mechanisms by which these 2 routes of antigen (Ag) delivery trigger CD4+ T cells during the priming and the recall phases. We will endeavor to uncover the cellular and molecular mechanisms that lead to effective or failed priming of T cells in the skin versus the lung. T cell receptor (TCR) transgenic (Tg) systems are arguably the best immunological tools to decipher the activation of clonal T cells in vivo, but there are no such reagents available for studying T-cell immunity to the dimorphic fungi. Therefore, we propose to create a Blastomyces-specific CD4+ TCR Tg mouse to monitor and analyze the activation of Ag-specific CD4+ T cells in these two immunologically disparate settings. We hypothesize that the subcutaneous delivery of Ag will prime TCR Tg, Blastomcyes-specific CD4+ T-cells in the skin-draining lymph nodes to become activated Th1 effector cells, and protect against lung challenge upon recall. Mucosal priming, on the other hand, will lead to the generation of functionally impaired Th1 effectors unable to provide resistance to challenge. Our specific aims are to: 1) Develop an adoptive transfer system to monitor Blastomyces-specific CD4+ T cells in vivo. Fungal-specific TCR Tg mice will be created from a T cell clone that protects mice against experimental infection. TCR Tg cells will be used to establish an autologous adoptive transfer system to track the phenotypic and functional properties of protective Ag-specific T cells during the priming phase in the skin, and on recall in the lung. 2) Identify whether and how mucosal priming fails to engender Ag-specific Th1 CD4+ cells. TCR Tg cells will let us track the critical stages of the CD4+ T cell response after Ag exposure at the respiratory mucosa: recruitment, activation, proliferation and differentiation in the lung and draining lymph nodes during the priming and recall phases. These events will be compared/contrasted to those after Ag exposure in the skin. Our studies will yield new insight into the evolution of T cell responses and immunity in mucosal tissues. This knowledge will contribute to the development of effective strategies to induce acquired, protective immunity for improved control of fungal and other microbial infections at these sites. PROJECT NARRATIVE: Fungal and other pathogens enter the body through the lung and cause disease at this site or after dissemination. The studies proposed here will create a uniquely powerful transgenic mouse to investigate the induction of protective immune responses in the lung. Our findings will provide the mechanistic basis to design strategies that prevent and/or treat infections with fungi and other microbial infections.