Deciphering the Pathogenesis of EHEC Infection and the Effects of Bacteria-Based Therapies Using Comparative Gut-on-a-Chip

使用比较芯片肠道破译 EHEC 感染的发病机制和细菌疗法的效果

• 批准号: 10612061
• 负责人: Yoko Miyamoto Ambrosini
• 金额: $ 22.98万
• 依托单位: WASHINGTON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10612061
• 关键词: Acute; Advanced Development; Alternative Therapies; Alzheimer' s Disease; Animal Model; Animal Testing; Antibiotic Therapy; Bacteria; Biochemical; Biological Assay; Biopsy; Canis familiaris; Cell Culture Techniques; Cells; Chronic; Clinical; Coculture Techniques; Colitis; Colonic Diseases; Colorectal Cancer; Coupling; Development; Diabetes Mellitus; Disease; Disease Progression; Epithelial Cells; Epithelium; Escherichia coli; Escherichia coli EHEC; Escherichia coli Infections; Experimental Designs; Exploratory/Developmental Grant for Diagnostic Cancer Imaging; Exposure to; Functional disorder; Future; Gastroenterology; Gastrointestinal Injury; Genomics; Germ-Free; Goals; Hemolytic-Uremic Syndrome; Hemorrhagic colitis; Human; Immune; Immune response; In Situ Hybridization; In Vitro; Individual; Infection; Intestines; Investigation; Lead; Maps; Medicine; Microfluidics; Mission; Modeling; Names; Neurologic; Organ; Outcome; Oxygen; Pathogenesis; Pathologic; Pathway interactions; Patients; Peripheral Blood Mononuclear Cell; Persons; Philosophy; Physiological; Physiology; Probiotics; RNA; Research; Resolution; Resources; Rodent; Rodent Model; Scientist; Shiga Toxin; Supportive care; System; Technology; Therapeutic; Therapeutic Effect; Therapeutic Intervention; Time; United States National Institutes of Health; Visualization; Work; canine model; combinatorial; comparative; cytotoxic; design; dysbiosis; foodborne; gut microbiome; high resolution imaging; high risk; human disease; in vitro Model; in vivo; insight; intestinal epithelium; intestinal homeostasis; metabolomics; microbiome; microcin; multiple omics; novel; novel therapeutics; organ on a chip; pathogen; physiologic model; preclinical development; prevent; spatiotemporal; stem cell biology; stem cells; transcriptomics

PROJECT SUMMARY This NIH ORIP R21 award application describes a 2-year plan designed to allow us to investigate pathophysiology of Enterohemorrhagic Escherichia coli (EHEC) infection in translational and comparative canine and human in vitro models. Investigation of the gastrointestinal injuries caused by EHEC and the development and assessment of therapeutic interventions have been hampered by the lack of translatable in vitro models that effectively reproduces the typical human colonic disease that progresses to bloody diarrhea and hemolytic uremic syndrome (HUS). The EHEC infection in dogs takes very similar clinical courses to that in humans as canine EHEC infection can naturally lead to mild to severe forms of EHEC infection as well as HUS. In carrying out the proposed research, development of both canine and human EHEC models will be accomplished while utilizing our expertise in veterinary gastroenterology, intestinal stem cell biology, microbiome, and microfluidic organ-on-chip technology. Our proposed study is a high-risk project because canine in vitro modeling of EHEC infection has never been performed before in an organ-chip platform. However, we believe our previous work makes this study highly feasible to develop canine and human EHEC Chips by incorporating colonoids, microbiome, immune cells, EHEC, and bacteria-based treatments into the physiological model. Specifically, Aim 1 will allow development of canine and human EHEC Chips and assessing the contribution of microbiome in the disease pathogenesis while mapping host responses by utilizing single-cell level multi-omics (especially genomics and transcriptomics) and RNA in situ hybridization. Aim 2. will allow assessment of immune contribution in the pathogenesis. Aim 3. will allow further assessment of preventative or therapeutic effects of two well studied bacteria (E.coli Nissle 1917 and MccPDI producing E. coli) on EHEC Chips. Consistent with the ORIP’s mission statements promoting veterinary scientists to employ their expertise in comparative medicine to investigate human diseases, my research will allow me to use my expertise in comparative gastroenterology as well as in primary stem cell culture to investigate alterations in intestinal homeostasis relevant to EHEC infection. The results generated in this proposal have direct implications for in vivo canine EHEC models and ultimately to human EHEC patients, since they will utilize donor-derived colonoids in the experimental designs and provide new insights into transcriptomic alterations in initiating, propagate, or ameliorate the EHEC infection. Our findings on the species similarities and differences in host responses can be applied to various chronic conditions that have been associated with intestinal dysbiosis that occur in both human and dogs (i.e., Colorectal Cancer, Diabetes Mellitus, and Alzheimer’s Disease, to name a few). In summary, the proposed study in this application will allow us to map the host-EHEC crosstalk as well as the effect of bacteria-based therapies by leveraging the single-cell multi-omics analysis, which may lead to a broader impact on uncovering the underlying disease mechanism and developing new preventative or therapeutic anti-EHEC therapy.

项目摘要 该NIH Orip R21奖申请描述了一项为期两年的计划，旨在使我们能够调查 转化和比较犬类中肠ha饮的大肠杆菌（EHEC）感染的病理生理 和人类体外模型。调查由EHEC造成的胃肠道损伤和发育 由于缺乏可翻译的体外模型，对治疗干预的评估受到了阻碍 有效地再现典型的人类结肠疾病，该疾病发展为血腥腹泻和溶血 尿毒症综合征（HUS）。狗的EHEC感染与人类的临床课程非常相似 犬EHEC感染自然会导致轻度至重度的EHEC感染和HUS。在携带中 将完成拟议的研究，犬类和人类EHEC模型的开发，同时 使用我们在兽医胃肠病学，肠道干细胞生物学，微生物组和微流体方面的专业知识 片上的器官技术。我们提出的研究是一个高风险项目，因为EHEC的犬类体外建模 从未在器官芯片平台上进行感染。但是，我们相信我们以前的工作 通过结合结肠素，使这项研究高度可行，可以开发犬类和人类EHEC芯片 微生物组，免疫细胞，EHEC和基于细菌的物理模型。具体来说，目标 1将允许开发犬和人EHEC芯片，并评估微生物组在 疾病发病机理在绘制宿主反应时使用单细胞水平多词（尤其是 基因组学和转录组学）和RNA原位杂交。目标2。将允许评估免疫 发病机理的贡献。 AIM 3。将允许进一​​步评估预防或治疗作用 在EHEC芯片上，两种良好的细菌（E.Coli Nissle 1917和McCPDI产生e.coli）。与 Orip的任务陈述促进兽医科学家在比较医学方面使用其专业知识 调查人类疾病，我的研究将使我能够在比较胃肠病学方面使用我的专业知识作为 以及在原发性干细胞培养中，以研究与EHEC感染相关的肠内稳态变化的改变。 本提案中产生的结果对体内犬EHEC模型有直接影响，最终对 人类EHEC患者，因为他们将在实验设计中利用供体衍生的结肠剂并提供 对启动，传播或改善EHEC感染的转录组改变的新见解。我们的发现 关于物种的相似性和宿主反应的差异，可以应用于各种慢性条件 与人和狗发生的肠道营养不良有关（即结直肠癌， 糖尿病和阿尔茨海默氏病，仅举几例）。总而言之，该应用中的拟议研究 将允许我们绘制主机-EHEC串扰以及基于细菌的疗法的效果。 单细胞多摩斯分析，这可能会导致对潜在疾病的更广泛影响 机制并开发新的预防或治疗性抗EHEC治疗。