GONOCOCCAL INFECTION AND GENE EXPRESSION IN FEMALE MICE

描述（改编自申请人摘要）：奈瑟氏球菌 淋病对妇女健康有严重影响， 这种病原体通过它感染上生殖道，而 导致严重的并发症（例如，慢性骨盆不自主疼痛 不孕症和异位妊娠）。目前的研究模式 淋球菌的发病机制是有限的能力，充分 模仿雌性生殖系统中宿主因素的复杂平衡 道。 因此，本建议的长期目标是 进一步建立一个雌性小鼠淋病模型，用于研究 适应N。淋球菌对宿主的时相和抗原性 表面分子的变化和体内基因表达。具体 为实现这一目标而设计的目标是：进一步表征 用于研究的雌二醇处理小鼠的实验感染 研究淋球菌生殖道具体方面的工具 感染; ii.确定在以下方面发挥作用的宿主因素： 选择淋球菌混浊（Opa）蛋白表达， 实验感染和测试的能力，Opa特异性 体内驱动Opa表型抗原性变异免疫应答; 三.鉴定实验过程中诱导的淋球菌基因 使用报告基因融合的鼠感染。拟议的实验 旨在实现这些目标的是：i.）雌二醇的敏感性- 将远交（SLC：ddY）和近交（BALB/6）小鼠用N.淋病 将根据感染的持续时间和程度进行表征 炎症。上生殖道感染将在 细菌与鼠子宫内膜相互作用的术语; ii.）主机 将研究在体内选择Opa阳性淋球菌的因素 通过监测Opa蛋白在嗜中性粒细胞耗尽小鼠中的表达， 补体缺陷小鼠和近交系小鼠， 炎症反应感染。PI将比较Opa 未免疫小鼠和小鼠阴道分离株的表型 用单一纯化的Opa蛋白抗原变体免疫， 确定Opa特异性免疫应答是否减少了 在体内表达同源Opa蛋白的淋球菌; iii.） 淋球菌过氧化氢酶-报告基因融合体的表达将被 在鼠感染期间和中性粒细胞粘附试验中测量， 研究淋球菌过氧化氢酶对炎症反应的调节。 利用绿色荧光报告基因构建转录基因融合库 将构建基因并筛选表达的启动子 在实验鼠感染期间;在这些下鉴定的基因 条件将被克隆用于进一步研究。

DESCRIPTION (Adapted from the applicant's abstract): Neisseria gonorrhoeae has a serious impact on women=s health due to the frequency with which this pathogen infects the upper reproductive tract, and the resultant serious complications (e.g., chronic pelvic pain, involuntary infertility and ectopic pregnancy). Current models for studying gonococcal pathogenesis are limited in their ability to sufficiently mimic the intricate balance of host factors in the female reproductive tract. Therefore, the long-term objectives of this proposal are to further develop a female mouse model of gonorrhea for studying the adaptation of N. gonorrhoeae to the host in terms of phase and antigenic variation of surface molecules and gene expression in vivo. The specific aims designed to achieve this objective are to: i. further characterize experimental infection of estradiol-treated mice for use as a research tool for studying specific aspects of gonococcal genital tract infection; ii. identify the host factor(s) that play a role in the selection for gonococcal opacity (Opa) protein expression during experimental infection and to test the capacity of an Opa-specific immune response to drive antigenic variation of Opa phenotype in vivo; iii. identify gonococcal genes that are induced during experimental murine infection using reporter gene fusions. The proposed experiments designed to address these aims are: i.) the susceptibility of estradiol- treated outbred (SLC:ddY) and inbred (BALB/6) mice to N. gonorrhoeae will be characterized with regard to duration of infection and degree of inflammation. Upper reproductive tract infection will be assessed in terms of bacterial interactions with the murine endometrium; ii.) host factors that select for Opa-positive gonococci in vivo will be studied by monitoring Opa protein expression in neutrophil-depleted mice, complement-deficient mice and inbred mice that uniformly do not produce inflammation in response to infection. The PI will compare the Opa phenotype of vaginal isolates from unimmunized mice and from mice immunized with a single purified Opa protein antigenic variant to determine if an Opa-specific immune response decreases the number of gonococci expressing the homologous Opa protein in vivo; iii.) expression of a gonococcal catalase-reporter gene fusion will be measured during murine infection and in neutrophil adherence assays to study the regulation of gonococcal catalase in response to inflammation. A transcriptional gene fusion bank using the green fluorescent reporter gene will be constructed and screened for promoters that are expressed during experimental murine infection; genes identified under these conditions will be cloned for further study.