Contribution of innate immune cells in promoting antibiotic tolerance

先天免疫细胞在促进抗生素耐受性方面的贡献

基本信息

批准号：
10300725
负责人：
Kim Davis
金额：
$ 24.56万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-06-01 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10300725
关键词：
Address Antibiotic Therapy Antibiotic susceptibility Antibiotics Bacteria Bacterial Genes Bacteriology Biological Models Cell Communication Cells Cellular Stress Complex Cues DNA Repair Development Diffuse Disease Distal Dose Doxycycline Environment Exhibits Exposure to Gene Expression Genes Growth Heterogeneity Imaging technology Immune Immune system Individual Infection Kinetics Knowledge Location Metabolic Metabolism Microbe Modeling Mus Nitric Oxide Pasteurella pseudotuberculosis Pathway interactions Patients Peripheral Phagocytes Phagocytosis Pharmacotherapy Phenotype Play Population Predisposition Process Production Proteome Reactive Nitrogen Species Reactive Oxygen Species Reporter Research Research Personnel Role Sepharose Spleen Stress System Systemic infection Testing Therapeutic Tissues Treatment Failure Type III Secretion System Pathway antibiotic tolerance antimicrobial bacterial community base biological adaptation to stress combat design extracellular human pathogen improved in vivo innovation insight macrophage metabolic rate mouse model neutrophil novel novel therapeutics response stressor tool transcriptome

项目摘要

PROJECT SUMMARY Despite high levels of antibiotic exposure, some individual bacterial cells survive multiple courses of antibiotic treatment due to antibiotic tolerance, which is a transient phenotypic change associated with reduced metabolism or slowed growth. It remains unclear what drives the formation of tolerant cells within host tissues, and whether the antimicrobials generated by the host immune system may contribute to the formation of antibiotic tolerant bacterial cells. To investigate the impact of host-derived stresses, specifically reactive oxygen species (ROS) and reactive nitrogen species (RNS), on antibiotic tolerance, we are probing the growth of the human pathogen, Yersinia pseudotuberculosis, in a mouse model. Within host tissues, this microbe replicates to form clonal clusters of extracellular bacteria that directly interface with a layer of neutrophils that are, in turn, enveloped by a layer of macrophages. The bacterial subpopulation at the outer edge of the microcolony responds to neutrophil contact by upregulating expression of the anti-phagocytic, type III secretion system (T3SS). These host cell-associated bacteria are part of a larger peripheral layer, which responds to and detoxifies diffusible nitric oxide (NO) originating from distal macrophages. This system clearly allows us to distinguish the spatial location of individual bacterial cells, and to determine if host cell interactions, or interactions with the antimicrobials they produce, are contributing to the formation of antibiotic tolerant bacterial populations. We hypothesize that antibiotic tolerant bacteria emerge within host tissues due to innate immune cell-derived ROS and RNS. We have recently developed a novel reporter system to detect antibiotic (doxycycline) exposure within individual bacterial cells in host tissues, and will use this tool to: 1) define the roles of immune cell-derived ROS and RNS in promoting of antibiotic tolerance, 2) determine the pathways utilized by doxycycline-tolerant subpopulations within host tissues. In addition, we have also developed an innovative agarose droplet-based system to model growth within host tissues, which will enable kinetic analyses, and provide a more thorough understanding of how bacterial communities interact with immune cells in vivo. The extent of bacterial heterogeneity within host tissues has been largely unexplored, making this a novel, understudied aspect of the disease process, and the focus of research in my lab. Identifying and understanding the cues that promote the formation of tolerant bacterial subpopulations has important implications for the design of novel therapeutics, which need to target all subpopulations to effectively eliminate bacteria within host tissues.

项目概要尽管接触高水平的抗生素，一些单个细菌细胞仍能在多个疗程中存活下来。由于抗生素耐受性而进行抗生素治疗，这是与减少相关的短暂表型变化新陈代谢或生长减慢。目前尚不清楚是什么驱动宿主组织内耐受细胞的形成，以及宿主免疫系统产生的抗菌剂是否可能有助于形成抗生素耐受的细菌细胞。研究宿主源性应激的影响，特别是反应性应激氧物种（ROS）和活性氮物种（RNS），关于抗生素耐受性，我们正在探索生长在小鼠模型中检测人类病原体假结核耶尔森氏菌。在宿主组织内，这种微生物复制形成细胞外细菌的克隆簇，直接与中性粒细胞层接触，反过来，又被一层巨噬细胞包围。外缘的细菌亚群微菌落通过上调抗吞噬细胞 III 型分泌物的表达来响应中性粒细胞接触系统（T3SS）。这些与宿主细胞相关的细菌是更大的外周层的一部分，该外周层对并解毒源自远端巨噬细胞的扩散性一氧化氮 (NO)。这个系统显然可以让我们区分单个细菌细胞的空间位置，并确定宿主细胞是否相互作用，或与其产生的抗菌剂相互作用，有助于形成抗生素耐受性细菌人口。我们假设，由于先天的原因，宿主组织内出现了抗生素耐受性细菌。免疫细胞衍生的 ROS 和 RNS。我们最近开发了一种新型报告系统来检测抗生素（多西环素）在宿主组织中单个细菌细胞内的暴露，并将使用此工具来：1）定义免疫细胞衍生的 ROS 和 RNS 在促进抗生素耐受中的作用，2) 确定途径由宿主组织内的多西环素耐受亚群利用。此外，我们还开发了一个基于创新的琼脂糖液滴的系统来模拟宿主组织内的生长，这将使得动力学分析，并更全面地了解细菌群落如何与免疫细胞相互作用体内。宿主组织内细菌异质性的程度在很大程度上尚未被探索，这使得这是一个疾病过程中新颖的、未被充分研究的方面，也是我实验室的研究重点。识别和了解促进耐受细菌亚群形成的线索具有重要意义对设计新疗法的影响，需要针对所有亚人群以有效消除宿主组织内的细菌。