Spotted Fever Rickettsial Antigens

描述(申请人提供)：严重斑点热立克次体病是一种威胁生命的、由壁虱传播的、新出现和再次出现的人类感染，由包括康氏立克次体和立克次体在内的专性细胞内立克次体引起。优秀的动物模型和现有的人类数据表明，干扰素-γ、CD8细胞毒性T淋巴细胞、树突状细胞(DC)和NK细胞在宿主保护性免疫中的关键作用。与自限性感染相比，在急性致死性小鼠斑点热立克次体病中，IL-10产生的CD4+CD25+T调节细胞显著抑制淋巴细胞增殖和IL-2和干扰素-γ的产生，这支持了人类立克次体感染相关免疫抑制的证据。目前尚不清楚为什么宿主防御系统未能控制致命立克次体病的细菌感染。这项研究的长期目标是更好地了解宿主防御系统无法控制重症斑点热立克次体感染所涉及的免疫调节机制。这项建议的目的是确定DC介导涉及T调节1细胞的先天缺陷和抑制的获得性免疫反应的机制，这可能导致致命的立克次体感染。我们的具体目标1是确定受损的DC-NK细胞串扰在促进严重斑点热立克次体缺陷的先天免疫反应方面的关键作用。我们将比较易感小鼠中有缺陷的DC-NK细胞串扰和耐药小鼠中有效的DC-NK细胞相互作用，前者会导致细菌负荷逐渐增加，后者导致先天反应中细菌的清除。我们的特异性目标2是确定免疫调节分子如PD-1/PD-L和吲哚胺2，3-双加氧酶或DC和/或T调节细胞表达或产生的细胞因子如IL-10在抑制重症斑点热立克次体T细胞应答中的作用。通过体内和体外实验，包括流式细胞术、ELISPOT、EL ISA、RT-或荧光定量聚合酶链式反应、小鼠TLR基因芯片、免疫组化染色、空斑分析、高效液相色谱检测等方法，我们将研究特异性目的1中介导DC-NK细胞相互作用的Toll样受体、细胞因子和趋化因子，以及DC与T调节细胞的相互作用在通过抑制性细胞因子和/或调节分子包括PD-1/PD-L和Ido介导T细胞和/或细胞凋亡中的作用。与公共卫生相关近年来，向疾控中心报告的落基山斑点热病例数量达到了历史最高水平。落基山斑热病是已知的最致命的立克次体疾病。该项目旨在确定免疫反应的哪些成分导致未能控制致命感染中的细菌生长，以使未来的免疫调节治疗能够提高存活率。

DESCRIPTION (provided by applicant): Severe spotted fever rickettsioses are life-threatening, tick-borne, emerging and re-emerging human infections caused by obligately intracellular Rickettsia including R. conorii and R. rickettsii. Excellent animal models and the available human data reveal the critical importance of IFN-gamma, CD8 cytotoxic T lymphocytes, dendritic cells (DCs) and NK cells in host protective immunity against Rickettsia. Evidence for human rickettsial infection-associated immunosuppression is supported by remarkable suppression of lymphocyte proliferation and IL-2 and IFN-gamma production by IL-10 producing CD4+CD25+ T regulatory cells in acute fatal murine spotted fever rickettsiosis compared to self-limited infection. It remains unclear why the host defense system fails to control bacterial infection in fatal rickettsiosis. The long-term goal of this research is to better understand the immune regulatory mechanisms involved in the inability of the host defense system to control infection in severe spotted fever rickettsiosis. The objective of this proposal is to determine the mechanisms by which DCs mediate defective innate and suppressed adaptive immune responses involving T regulatory 1 cells, which may lead to fatal rickettsial infection. Our specific aim 1 is to determine the key effect of impaired DC-NK cell cross talk on promoting a defective innate immune response in severe spotted fever rickettsiosis. We will compare the defective DC-NK cell cross talk in susceptible mice, which causes progressively increased bacterial loads, with efficient DC-NK cell interaction in resistant mice, which lead to clearance of bacteria during the innate response. Our specific aim 2 is to determine the role of immunoregulatory molecules such as PD-1/PD-L and indoleamine 2,3-dioxygenase (IDO) or cytokines such as IL-10 expressed or produced by DCs and/or T regulatory cells in suppression of T cell responses during severe spotted fever rickettsiosis. Toll-like receptors (TLRs), cytokines and chemokines that mediate the DC-NK cell interaction in specific aim 1, and the role of interactions of DCs with T regulatory cells in mediating suppressed protective effector type-1 T cells and/or apoptosis via suppressive cytokines and/or regulatory molecules including PD-1/PD-L and IDO in severe spotted fever rickettsiosis in specific aim 2 will be investigated using in vivo and in vitro approaches including flow cytometry, ELISPOT, ELISA, RT- or real time PCR, mouse TLR PCR array, immunohistochemical staining, plaque assay, HPLC as well as adoptive transfer, depletion of immune molecules and knockout mice. PUBLIC HEALTH RELEVANCE In recent years the number of cases of Rocky Mountain spotted fever, the most lethal rickettsial disease known, reported to the CDC has reached the highest level in history. This project is designed to determine what components of the immune response are responsible for failure to control bacterial growth in fatal infections to allow future immunomodulatory treatment to enhance survival.