Cooperation of CD8+ T cells and phagocytes to eliminate Toxoplasma cysts

描述（由申请人提供）：弓形虫是一种重要的慢性感染，能够在艾滋病患者和其他免疫功能低下的个体中引起危及生命的弓形虫脑炎。感染持续存在的基础是组织囊肿，它在宿主的生命中基本上保持静止，但可以重新激活并引起疾病。这个阶段的寄生虫不受任何现有药物治疗的影响，通常被认为是不可触及的。然而，我们最近的研究表明，免疫T细胞能够消除T。当T细胞被转移到已经产生大量囊肿的受感染免疫缺陷动物体内时，这种T细胞介导的免疫过程与组织囊肿周围单核细胞（主要是形态学上的小胶质细胞和巨噬细胞）的积累有关。由于积聚的吞噬细胞渗透到囊肿内，这些细胞似乎是破坏囊肿的主要效应细胞，并在免疫T细胞启动该过程后将其从大脑中消除。CD 8+免疫T细胞具有启动这种抗囊肿免疫过程的有效活性，并且T细胞的保护活性需要穿孔素。总目标 本研究的目的是探讨穿孔素介导的CD 8+免疫T细胞协同吞噬细胞清除T细胞的分子机制。从大脑中取出弓形虫囊肿在第一个目标中，我们将确定CD 8+免疫T细胞是否积聚并附着在大脑中的含囊肿细胞上。我们还将确定CD 8 + T细胞是否在体外以穿孔素依赖性方式裂解含囊细胞，以及CD 8 + T细胞的穿孔素介导的活性是否诱导吞噬细胞在脑中囊周围的积累。在第二个目标中，我们将确定对CD 8 + T细胞识别含囊细胞至关重要的MHC I类分子。然后我们将确定T。通过筛选预测的缓殖子表位肽和/或缓殖子cDNA文库，由鉴定的MHC I类分子呈递的弓形虫包囊表位，用于被CD 8 + T细胞识别。第三个目的是确定吞噬细胞清除T。弓形虫囊肿后，启动免疫过程的CD 8 + T细胞。我们将研究诱导型一氧化氮合酶和自噬在杀死缓殖子和消除囊肿中的作用，使用缺乏每种机制所需的分子的小鼠。从这三个特定目标的研究中产生的信息将为开发激活CD 8 + T细胞和吞噬细胞以消除T细胞的新方法提供必要的基础。弓形虫囊肿可以重新激活并导致弓形虫脑炎。

DESCRIPTION (provided by applicant): Toxoplasma gondii establishes an important chronic infection capable of causing life-threatening toxoplasmic encephalitis in AIDS patients and other immunocompromised individuals. The basis of persistence of the infection is the tissue cyst, which remains largely quiescent for the life of the host, but can reactivate and cause disease. This stage of the parasite is not affected by any of the current drug treatments and it has been generally regarded as untouchable. However, our recent studies revealed that the immune T cells are able to eliminate T. gondii cysts from the brains of infected hosts when the T cells are transferred into infected immunodeficient animals that have already developed large numbers of the cysts. This T cell-mediated immune process is associated with accumulation of mononuclear cells, primarily microglia and macrophages morphologically, around tissue cysts. Since the accumulated phagocytes penetrate within the cyst, these cells appear to be the main effector cells that destroy the cysts and eliminate them from the brain after initiation of this process by immune T cells. CD8+ immune T cells possess a potent activity to initiate this anti-cyst immune process, and the protective activity of the T cells requires perforin. The overall goal of this research project is to determine the molecular mechanisms by which perforin-mediated activity of CD8+ immune T cells cooperates with the phagocytes to remove T. gondii cysts from the brain. In the first aim, we will determine if CD8+ immune T cells accumulate and attach to the cyst-containing cells in the brain. We will also determine if the CD8+ T cells lyse cyst-containing cells in a perforin-dependent manner in vitro and whether the perforin- mediated activity of CD8+ T cells induces the accumulation of phagocytes around the cysts in the brain. In the second aim, we will determine the MHC class I molecule(s) critical for recognition of cyst-containing cells by CD8+ T cells. We will then determine the T. gondii cyst epitope(s) presented by the identified MHC class I molecule for recognition by the CD8+ T cells by screening predicted bradyzoite epitope peptides and/or bradyzoite cDNA library. The third aim is to determine the mechanisms by which phagocytes eliminate T. gondii cysts after initiation of the immune process by CD8+ T cells. We will examine the roles of inducible nitric oxide synthase and autophagy in killing of bradyzoites and elimination of cysts using mice lacking a molecule required for each mechanism. The information generated from the studies in these three specific aims all together will provide the essential basis for developing a novel method(s) to activate CD8+ T cells and phagocytes to eliminate T. gondii cysts that can reactivate and cause toxoplasmic encephalitis.