Deciphering the source and consequences of Nur77 expression during E. coli rUTI in mice

摘要 复发性尿路感染（鲁蒂）是一个昂贵的临床问题，影响了全球数百万妇女， 导致全球抗生素耐药性上升。多种证据表明阴道中的细菌 会影响UTI的易感性。例如，患有阴道微生态失调的女性称为细菌性阴道病（BV） 患UTI的风险增加。几种阴道细菌-包括BV相关物种阴道加德纳菌 （G.v.）─通过针吸或经尿道直接从膀胱收集的尿液中分离 漂浮导管尽管培养条件通常不足以检测到它，但尿液中G. v的存在也是 在流行病学上与鲁蒂有关。我们已经开发了新的小鼠模型来定义阴道给药的影响。 从一个新的角度测试的想法，短暂的膀胱暴露于阴道细菌对尿路感染 如G. v.影响大肠杆菌（E.c.）的致病性，UTI的主要原因。我们之前 表明G. v.触发小鼠膀胱上皮细胞凋亡和脱落。当老鼠窝藏 潜伏性膀胱内贮器从先前的实验感染中，G. v.引发了E. c的出现。 和鲁蒂的速率比暴露于单独媒介物的小鼠高4倍。这些数据表明膀胱暴露 可能是女性鲁蒂的诱因 为了在细胞和分子水平上了解G. v.如何触发脱落和鲁蒂，我们检查了 在膀胱中使用批量RNA测序对G. v.的转录响应。这一分析表明，G. V. 诱导孤儿核受体Nur 77（也称为Nr 4a 1）的表达。Nur 77是公认的 它在细胞凋亡和其他组织疾病的炎症反应中的作用，但它在泌尿道中的作用是 大部分未经研究。我们使用Nur 77-/-小鼠的初步数据表明，Nur 77是G. v.触发E.c. 鲁蒂在我们的小鼠模型中。在这篇聚焦R 03的文章中，我们试图回答两个关于 Nur 77对G. v.的贡献-诱导E.c.鲁蒂：G. v.后膀胱中哪些细胞类型表达Nur 77。 曝光？(Aim 1），以及宿主对G. v.脱落或炎性细胞因子的反应 Nur 77介导的生产？(Aim 2）。我们提出的实验建立在我们的初步数据和 利用现有的Nur 77敲除和报告小鼠以及我们在详细描述膀胱反应方面的经验， 微生物暴露。这些研究的成功完成将提供重要的机理见解， G. v.触发E.c.鲁蒂，并将生成关键数据，在此基础上构建未来的R 01提案。

ABSTRACT Recurrent urinary tract infection (rUTI) is a costly clinical problem that affects millions of women worldwide and contributes to the global rise in antibiotic resistance. Multiple lines of evidence indicate that bacteria in the vagina can affect susceptibility to UTI. For example, women with the vaginal dysbiosis called bacterial vaginosis (BV) are at increased risk of UTI. Several vaginal bacteria—including the BV-associated species Gardnerella vaginalis (G.v.)—have been isolated from urine collected directly from the bladder by needle aspiration or transurethral catheterization. Although culture conditions are not often poised to detect it, the presence of G.v. in urine is also epidemiologically linked with rUTI. We have developed new mouse models to define the impact of vaginal bacteria on UTI from a new perspective—testing the idea that transient bladder exposures to vaginal bacteria such as G.v. influence the pathogenesis of Escherichia coli (E.c.), the leading cause of UTI. We previously demonstrated that G.v. triggers apoptosis and exfoliation of the bladder epithelium in mice. When mice harbored latent intracellular bladder reservoirs of E.c. from a prior experimental infection, G.v. triggered emergence of E.c. and rUTI at a rate 4-fold higher than mice exposed to vehicle alone. These data indicate that bladder exposures to G.v. could represent a trigger of rUTI in women. To understand at the cellular and molecular level how G.v. triggers exfoliation and rUTI, we have examined the transcriptional response to G.v. in the bladder using bulk RNA sequencing. This analysis revealed that G.v. induced the expression of the orphan nuclear receptor Nur77 (also called Nr4a1). Nur77 is well-recognized for its role in apoptosis and in inflammatory responses in diseases in other tissues, but its role in the urinary tract is largely unstudied. Our preliminary data using Nur77-/- mice indicate that Nur77 is required for G.v. to trigger E.c. rUTI in our mouse model. In this focused R03, we seek to answer two fundamental questions regarding the contribution of Nur77 to G.v.-induced E.c. rUTI: Which cell types in the bladder express Nur77 following G.v. exposure? (Aim 1), and Which host responses to G.v. exposure—exfoliation or inflammatory cytokine production—are mediated by Nur77? (Aim 2). Our proposed experiments build upon our preliminary data and leverage available Nur77 knockout and reporter mice and our experience in detailing bladder responses to microbial exposures. Successful completion of these studies will provide important mechanistic insights into how G.v. triggers E.c. rUTI and will generate key data on which to build a future R01 proposal.