Mycobacterial antigen compartmentalization & immunity

结核病仍然是现代世界面临的最重大的公共卫生挑战之一。 需要改进的疫苗来预防感染，需要改进的免疫疗法来对抗现有或复发的疾病。 更好的疫苗将通过CD4+和CD8+ T细胞一起诱导协调的保护性应答。 我们的假设是CD4+和CD8+ T细胞对不同的细菌隔室进行不同的采样。关于哪个区室对于有效呈递II类和I类表位和产生保护性应答是最佳的信息将有助于创造更好的疫苗。 为了做到这一点，我们将使用一种新的方法，同时建立在以前的实验系统的优势。 TCR转基因小鼠将感染重组牛分枝杆菌菌株卡介苗（BCG），目前的疫苗株。 这些rBCG将在位于细菌的不同亚细胞区室中的相同融合蛋白的背景下表达T细胞表位。 将分别使用脉络丛脑膜炎病毒（LCMV）gp33肽或鸽子细胞色素C肽（PCC）构建I类或II类呈递的rBCG菌株的平行系列。 我们将研究每个表位进入其各自的呈递途径如何受到其在不同细菌隔室中的位置的影响。随后，我们将使用过继转移的抗原特异性T细胞研究抗原特异性细胞的激活和募集，包括全身性的和BCG诱导的肝肉芽肿中的抗原特异性细胞对各种rBCG的应答。 我们的最终分析将是研究当抗原在不同的细菌隔室中呈递时，在有或没有预先肽特异性免疫的情况下，菌血症是如何受到影响的。 通过这种方式，我们希望确定不同细菌区室中的不同表位如何影响T细胞应答和保护。 我们选择PCC（CD4+特异性）和gp33（CD8+特异性）进行这项工作，因为它们都是广泛研究的模型抗原，并且有多种试剂可用，包括T细胞克隆，杂交瘤，TCR转基因小鼠和MHC四聚体试剂。 选择BCG感染的小鼠模型是因为我们希望提高该减毒株的疫苗能力，还因为用BCG感染小鼠已被广泛使用，并且该模型的许多特征已被充分理解。 该提案的实验结果应该与改进预防结核病的疫苗设计直接相关，并且还将提供有关细菌抗原如何进入不同抗原呈递途径的知识。

Tuberculosis remains one of the most significant public health challenges the modern world faces. Improved vaccines are needed for prevention of infection and improved immunotherapies are needed to combat existing or recurring disease. An better vaccine would induce a concerted protective response by CD4+ and CD8+ T cells together. Our hypothesis is that CD4+ and CD8+ T cells sample different bacterial compartments differently. Information about which compartment is optimal for effective presentation of ClassII and Class I epitopes and generation of protective responses will help create better vaccines. To do this we will use a novel approach while building on the strengths of previous experimental systems. TCR transgenic mice will be infected with recombinant Mycobacterium bovis strain bacille Calmette Guerin (BCG), the current vaccine strain. These rBCG will express T cell epitopes in the context of the same fusion proteins located in different subcellular compartments of the bacteria. A parallel series of rBCG strains will be constructed for both class I or class II presentation using either Lymphochoriomeningitis Virus (LCMV) gp33 peptide or pigeon cytochrome C peptide (PCC) respectively. We will study how access of each epitope to its respective presentation pathway is influenced by its location in different bacterial compartments. Subsequently, we will study the activation and recruitment of antigen specific cells both systemically and in the BCG induced liver granulomas in response to various rBCG using adoptively transferred antigen specific T cells. Our final analysis will be to study how bacteremia is effected when antigen is presented in different bacterial compartments with or without prior peptide specific immunization. In this manner we hope to define how the different epitopes in different bacterial compartments effect T cell responses and protection. We chose PCC (CD4+ specific) and gp33 (CD8+ specific) for this work because they are both widely studied model antigens and a multitude of reagents are available, including T cell clones, hybridomas, TCR transgenic mice, and MHC tetramer reagents. The mouse model of BCG infection was chosen because we wish to improve the vaccine capacity of this attenuated strain and also because infection of mice with BCG has been widely employed and many of the characteristics of this model are well understood. The experimental results from this proposal should have direct relevance to improving vaccine design for protection against tuberculosis, and will also provide knowledge about how bacterial antigen access different antigen presenting pathways.