Identifying NKT cell lipids of Chlamydia trachomatis and C. muridarum

描述(由申请人提供)：沙眼衣原体是最常见的可报告的性传播感染(STI)，每年导致美国100万例和全球约9200万例。生殖器感染可导致女性生殖器官免疫介导的损害和严重的生殖障碍，包括盆腔炎(PID)，可导致慢性盆腔疼痛、宫外孕和不孕症。每年大约有8%的女性患上产后窘迫症，在再次感染后，这种风险会显著增加40%-70%。再感染率约为13%，发生在6个月内。从经济角度来看，衣原体感染的治疗和肺炎的治疗每年花费美国医疗保健系统数十亿美元。该项目调查了导致产后妊娠综合征/不孕症的潜在因素，其结果可用于 用于开发治疗方法，以预防产后不孕症/不孕，并最终降低治疗成本。沙眼衣原体生殖道感染小鼠模型被用来揭示不孕症/输卵管性不孕的潜在机制(S)，被称为上生殖道病理学(UGTP)。我们的体外和体内研究表明，如其他人所证实的那样，iNKT可以通过感染鼠疫杆菌而被激活。众所周知，iNKT细胞调节免疫应答的结果，并可能在针对衣原体的免疫应答中发挥作用。为了了解iNKT细胞如何调节抗衣原体免疫，即帮助辐射或导致女性生殖道功能障碍，需要识别CD1d结合的糖脂。我们以前的数据显示，衣原体中含有的糖脂抗原在体外用无细胞CD1d抗原提呈试验激活了I型(INKT)和II型NKT细胞。一种II型微生物抗原尚未在任何微生物中被鉴定，这使得衣原体在NKT细胞生物学研究中是独一无二的。脂类对专性细胞内衣原体的生存非常重要，衣原体从宿主细胞合成一些脂类，并将其他脂类结合到细胞壁中。CD1d降解是这些细菌免疫逃避的一种策略，这并不令人惊讶。具体目标1，我们建议与分离CD1d结合糖脂的专家威廉·希尔德布兰德博士合作，鉴定衣原体中包含的激活NKT细胞的CD1d结合抗原。在特定的目标2中，我们将验证哪些糖脂激活 通过对衣原体生殖器感染后缺乏iNKT细胞(C57BL/6 Bkgd上的CD1d-/-)的小鼠匀浆组织进行流式细胞术和ELISA检测，通过检测体内的细菌负荷、Th1细胞数量、细胞因子和趋化因子分泌以及UGtP来说明激活的iNKT细胞对衣原体生殖道感染结局的调节程度。了解iNKT细胞如何影响衣原体生殖器感染是NKT细胞生物学的一个重要问题，将与NKT细胞生物学专家Mitchell Kronenberg博士合作进行研究。

DESCRIPTION (provided by applicant): C. trachomatis is the most common reportable sexually transmitted infection (STI) and is responsible for > 1 million cases in the U.S. and approximately 92 million cases worldwide each year. Genital infection can lead to immune-mediated damage of the female reproductive organs and serious reproductive disability, including pelvic inflammatory disease (PID) that can result in chronic pelvic pain, ectopic pregnancy and infertility. Approximately 8% of females annually develop PID and this risk significantly increases by 40-70% following re- infection. The reinfection rate is approximately 13% and occurs within 6 months. Viewed in economic terms, treatment of chlamydial STIs and treatment of PID costs the US health care system billions of dollars annually. This project examines a potential contributing factor for developing PID/infertility and the results can be used for development of therapeutics to prevent PID/infertility and ultimately reduce treatment costs. The mouse model of C. trachomatis genital infection (C. muridarum) is used to reveal the underlying mechanism(s) for developing PID/tubal infertility and is called upper genital tract pathology (UGTP). Our in vitro and in vivo studies show that iNKT are activated by infection with C. muridarum as others have confirmed. It is well known that iNKT cells modulate the outcome of immune responses and likely play a role in the immune response against Chlamydia. In order to understand how iNKT cells modulate anti-chlamydial immune, ie aid in eradiation or contribute to female reproductive tract dysfunction, the identity of CD1d-binding glycolipids are needed. Our previous data revealed that glycolipid antigens contained within chlamydiae activate type I (iNKT) and type II NKT cells in vitro using a cell-free CD1d antigen presentation assay. A type II microbial antigen has not yet been identified for any microbe making Chlamydia unique for the study of NKT cell biology. Lipids are important for survival of the obligate intracellular chlamydiae which synthesize a few lipids and incorporate others into their cell wall from host cells. It is not surprising that CD1d degradation is a strategy for immune evasion of these bacteria. Specific Aim 1, we propose to identify CD1d-binding antigens contained within chlamydiae that activate NKT cells in collaboration with Dr. William Hildebrand, an expert in isolating CD1d- binding glycolipids. In Specific Aim 2, we will validate which glycolipids activate iNKT cells in vivo and demonstrate the extent to which activated iNKT cells modulate the outcome of chlamydial genital infection by measuring bacterial burden, Th1 cell numbers, cytokine and chemokine secretion and UGTP in vivo using flow cytometry and ELISA assays on homogenized tissue from mice that lack iNKT cells (CD1d-/- on C57BL/6 Bkgd) after chlamydial genital infection. Understanding how iNKT cells influence chlamydial genital infection is an important question of NKT cell biology and will be studied in collaboration with Dr. Mitchell Kronenberg, an expert in NKT cell biology.