Microbiota based mechanisms of post-infection irritable bowel syndrome

感染后肠易激综合征的微生物群机制

• 批准号: 10364382
• 负责人: Madhusudan Grover
• 金额: $ 34.98万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-21 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10364382
• 关键词: Acute; Adult; Affect; Automobile Driving; Bacteria; Bacteroides; Beta-glucuronidase; Bilirubin; Biological Markers; Campylobacter jejuni; Chronic; Communities; Coupled; Data; DegP protease; Development; Diarrhea; Engraftment; Enterocolitis; Enzymes; Escherichia coli; Family; Feces; Functional disorder; Funding; Germ-Free; Health; Healthcare; Human; Human Activities; Impairment; In Vitro; Infection; Intervention; Intestines; Irritable Bowel Syndrome; Link; Longitudinal Studies; Mediating; Mediator of activation protein; Metabolism; Metagenomics; Microbe; Morbidity - disease rate; Mus; Pathway interactions; Patients; Peptide Hydrolases; Play; Population; Prevotella; Protease Inhibitor; Proteins; Regulation; Regulation of Proteolysis; Risk; Risk Factors; Role; Sampling; Serine Protease; Serine Proteinase Inhibitors; Source; Structure; Symptoms; Testing; Tight Junctions; Time; Transplantation; base; commensal microbes; disabling symptom; dysbiosis; enteric infection; experimental study; fecal microbiota; gut bacteria; gut microbiota; human microbiota; humanized mouse; improved; in vivo; inhibitor/antagonist; intestinal barrier; metaproteomics; microbial; microbial community; microbiota; mouse model; overexpression; restoration; selective expression

ABSTRACT One in six adults in the U.S. suffers from chronic and often disabling symptoms of irritable bowel syndrome (IBS). Intestinal infections are an established risk-factor for development of post-infection IBS (PI-IBS). The intestinal tract contains a variety of proteases, and we have discovered that PI-IBS patients have significantly higher fecal proteolytic activity (PA) than controls. More importantly, PA associates strongly with loss of intestinal barrier function and worse symptoms for the patients. We found that patients who develop PI-IBS and have high PA have a significant loss of microbial diversity that starts soon after the infection. Key microbial taxa are lost, especially from the Alistipes genus. Using metaproteomics, we found that proteases driving PA in these patients are of human origin. In order to understand if loss of microbiota could be affecting host proteases, we used germ-free mice. Colonization of germ-free mice with healthy human microbiota (humanization) results in a significant decline of PA suggesting commensal microbes inhibit host proteases and thus have a role in maintaining intestinal health. However, the dysbiotic microbiota from the high PA patients that are missing specific microbes were unable to suppress PA. We hypothesize that Alistipes and other missing bacteria play a critical role in suppression of PA. We plan to test the candidate bacteria identified in the preliminary experiments and inter-species interactions in PA regulation in Aim 1. Next, we determined how loss of microbes result in poor inhibition of proteases. Unconjugated bilirubin is an inhibitor of serine proteases and microbial β-glucuronidases deconjugate bilirubin. We found that the PI-IBS patients with high PA have lower fecal microbial β-glucuronidase enzymatic activity. Additionally, they have lower levels of end products of bilirubin deconjugation. β-glucuronidases are a large family of microbial enzymes with varying sources, structures and catalytic efficacies for different substrates. We hypothesize that loss of specific microbial β-glucuronidases will result in impaired deconjugation of bilirubin. In Aim 2, we will analyze metagenomics data from our PI-IBS patients with high and low PA for presence of microbial β-glucuronidases as well as determine the efficacy of these fecal samples for bilirubin deconjugation. Additionally, we will generate purified β-glucuronidases from Alistipes and other bacterial taxa for assessing bilirubin deconjugation efficacy in vitro. Next, we have shown that fecal microbiota transfer using an Alistipes enriched low PA community can suppress PA in high PA humanized mice providing a rationale for using microbiota for protease suppression and correcting intestinal barrier function. In Aim 3, we will use cohousing strategies to allow microbiota transfer between humanized high and low PA mice and determine if barrier dysfunction associated with a high PA state can be reversed. Furthermore, we will determine changes in barrier pathways, ionic selectivity and expression of tight junction proteins upon engraftment of new microbiota. Together, these aims will examine microbial influence on PI-IBS pathophysiology via regulation of intestinal proteases. Identification of microbiota-based strategies that can result in protease inhibition and restoration of barrier function can be beneficial for IBS and other conditions associated with microbial dysbiosis.

抽象的 在美国，六分之一的成年人患有慢性且常常致残的肠易激综合症症状 （肠易激综合症）。肠道感染是感染后 IBS (PI-IBS) 发展的既定危险因素。这 肠道内含有多种蛋白酶，我们发现PI-IBS患者有明显的 粪便蛋白水解活性（PA）高于对照组。更重要的是，PA 与丧失 肠道屏障功能受损，患者症状恶化。我们发现，患有 PI-IBS 的患者 PA 较高的细菌在感染后不久就会出现微生物多样性的显着丧失。关键微生物 类群消失，尤其是阿里斯提属 (Alistipes)。使用宏蛋白质组学，我们发现驱动 PA 的蛋白酶 这些患者是人类。为了了解微生物群的丧失是否会影响宿主 蛋白酶，我们使用无菌小鼠。健康人类微生物群在无菌小鼠中的定植 （人源化）导致 PA 显着下降，表明共生微生物抑制宿主蛋白酶 从而具有维持肠道健康的作用。然而，高 PA 导致的菌群失调 缺少特定微生物的患者无法抑制 PA。我们假设 Alitipes 和 其他缺失的细菌在抑制 PA 中发挥着关键作用。我们计划测试已确定的候选细菌 在目标 1 中 PA 调节的初步实验和物种间相互作用中。接下来，我们确定 微生物的损失如何导致蛋白酶抑制效果不佳。非结合胆红素是丝氨酸的抑制剂 蛋白酶和微生物 β-葡萄糖醛酸酶可解离胆红素。我们发现 PI-IBS 患者的高 PA具有较低的粪便微生物β-葡萄糖醛酸酶活性。此外，它们的末端水平较低 胆红素解离产物。 β-葡萄糖醛酸酶是一个微生物酶大家族，具有不同的性质 不同底物的来源、结构和催化效率。我们假设特定的损失 微生物β-葡萄糖醛酸酶会导致胆红素解离受损。在目标 2 中，我们将分析 来自高 PA 和低 PA 的 PI-IBS 患者的微生物 β-葡萄糖醛酸酶存在的宏基因组数据 以及确定这些粪便样本对胆红素解离的功效。此外，我们将 从 Alistipes 和其他细菌分类群中产生纯化的 β-葡萄糖醛酸酶，用于评估胆红素解结合 体外疗效。接下来，我们展示了使用 Alistipes 富集低 PA 进行粪便微生物群转移 群落可以抑制高 PA 人源化小鼠中的 PA，为使用微生物群作为蛋白酶提供了理论依据 抑制和纠正肠道屏障功能。在目标 3 中，我们将使用共同居住策略来允许 人源化高 PA 小鼠和低 PA 小鼠之间的微生物群转移，并确定屏障功能障碍是否相关 高PA状态可以逆转。此外，我们将确定屏障途径、离子通道的变化 新微生物群植入后紧密连接蛋白的选择性和表达。这些目标共同实现 将通过调节肠道蛋白酶来检查微生物对 PI-IBS 病理生理学的影响。鉴别 基于微生物群的策略可以抑制蛋白酶并恢复屏障功能 对肠易激综合症和其他与微生物失调相关的疾病有益。