Modulation of Host-Microbe Interactions by Extracellular Purines in the Gastrointestinal Lumen

项目摘要/摘要 在美国，超过300万人被诊断为炎症性肠病(IBD)，这是一种慢性疾病 以及原因不明的不治之症，伴随着巨大的发病率和医疗费用。 肠道生物失调是IBD的一个标志，尽管有特定的机制，但它在其发病机制中起到了作用。 这一观察的基础是缺乏的。作为一名胃肠病专家和内科科学家，候选人寻求 阐明介导宿主-微生物相互作用的特定分子机制，从而推动IBD与终极 将这些发现转化为临床应用的目标。这项拟议的研究建立在候选人的 腺苷(ADO)调节肠道沙门氏菌定植和毒力的已发表观察 并将为候选人提供必要的额外培训，使其成为一名 独立调查员。然而，已有研究表明，胃肠道粘膜中的腺苷信号可促进分辨率 在炎症方面，尚未研究腺苷对肠道微生物区系的作用。建议数 研究验证了胃肠道粘膜产生的细胞外ADO影响 肠道微生物区系的组成和代谢通过抑制细菌的应激反应 (Sr)抑制生物失调，促进肠道内环境平衡。这一假设是通过三个目标来探讨的： 目的1、明确腺苷抑制肠道细菌生长的机制。初步数据显示 ADO通过抑制SR而诱导氨基酸营养缺乏症，从而抑制细菌的生长。 细菌对营养压力的适应。这一观察结果背后的机制将被确定 通过对缺乏嘌呤代谢的细菌突变体进行代谢学和转录组分析。目标2将 阐明腺苷通过调节肠道功能促进肠道内环境平衡的机制 微生物区系。一种新的肠道上皮细胞缺乏细胞外腺苷生成能力的小鼠模型 (条件敲除ECTO-5‘-核苷酸酶)将被用来评估ADO如何塑造肠道微生物区系。 小鼠粪便微生物转移(FMT)-IBD将用于检测ADO在促进肠道功能方面的作用 通过塑造肠道微生物区系实现动态平衡。最后，Aim 3展示了 正在研究的机制表明，外源性注射的嘌呤可以改变肠道 以类似于肠道中自然产生的嘌呤的方式的动态平衡。拟议的工作将是 在哥伦比亚大学肖恩·科尔根博士和安德烈·S·巴斯克斯-托雷斯博士的指导下演出 科罗拉多州，他们在腺苷在胃肠道炎症中的作用和 核苷酸代谢在肠道传染病发病机制中的作用。这个指导团队是 非常适合于拟议的研究，该研究位于他们每个专业领域的交叉点 并将在IBD小鼠模型领域为候选人提供必要的额外培训， 代谢分析、微生物学和微生物组分析。

Project Summary/Abstract Over 3 million people in the United States carry a diagnosis of inflammatory bowel disease (IBD), a chronic and incurable condition of unknown cause that carries with it significant morbidity and healthcare costs. Intestinal dysbiosis is a hallmark of IBD that contributes to its pathogenesis, though specific mechanisms underlying this observation are lacking. As a gastroenterologist and physician scientist, the candidate seeks to elucidate specific molecular mechanisms that mediate host-microbe interactions that drive IBD with the ultimate goal of translating these findings into clinical applications. The proposed research builds on the candidate's published observations that adenosine (Ado) regulates colonization and virulence of the Salmonella enterica serovar Tyhpimurium and will provide the candidate with the additional training necessary to move forward as an independent investigator. Whereas it has been shown that Ado signaling in the GI mucosa promotes resolution of inflammation, the actions of Ado on the intestinal microbiota have not been examined. The proposed research tests the hypothesis that extracellular Ado produced in the gastrointestinal mucosa influences the composition and metabolism of the intestinal microbiota through inhibition of the bacterial stringent response (SR) to inhibit dysbiosis and promote intestinal homeostasis. This hypothesis is explored through three aims: In Aim 1, the mechanism by which Ado inhibits growth of enteric bacteria will be defined. Preliminary data show that Ado inhibits bacterial growth by inducing amino acid auxotrophies through suppression of the SR, a global bacterial adaptation to nutritional stress. The mechanisms underlying this observation will be determined through metabolomic and transcriptomic analyses of bacterial mutants deficient in purine metabolism. Aim 2 will elucidate the mechanisms by which Ado promotes intestinal homeostasis by modulating the intestinal microbiota. A novel mouse model wherein intestinal epithelial cells lack the ability to generate extracellular Ado (conditional knockout of ecto-5'-nucleotidase) will be used to assess how Ado shapes the intestinal microbiota. Murine fecal microbial transfer (FMT)-IBD will be used to examine the role of Ado in promoting intestinal homeostasis by shaping the intestinal microbiota. Finally, Aim 3 presents a translational application of the mechanisms under study by demonstrating that exogenously administered purines can alter intestinal homeostasis in an analogous manner to naturally-occurring purines in the intestine. The proposed work will be performed under the mentorship of Drs. Sean Colgan and Andrés Vázquez-Torres at the University of Colorado, who have extensive expertise in the role of adenosine in gastrointestinal inflammation and the role of nucleotide metabolism in the pathogenesis of enteric infectious diseases, respectively. This mentoring team is ideally suited to the proposed research, which is situated at the intersection of each of their areas of expertise and will provide the candidate with necessary additional training in the areas of murine models of IBD, metabolomic analysis, microbiology, and microbiome analysis.