Pomegranate Extract and Its Microbial Metabolite Urolithin A Suppress IBD through Modulation of the Gut Microbiome and T Cell Inflammatory Immune Responses

石榴提取物及其微生物代谢物尿石素 A 通过调节肠道微生物群和 T 细胞炎症免疫反应来抑制 IBD

• 批准号: 10363573
• 负责人: Zhaoping Li
• 金额: --
• 依托单位: VA GREATER LOS ANGELES HEALTHCARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10363573
• 关键词: Adjuvant Therapy; Affect; Alternative Therapies; Anti-Inflammatory Agents; Antiinflammatory Effect; Attenuated; Bacteria; Blood; CD4 Positive T Lymphocytes; Cell Line; Cells; Chemicals; Chronic; Chronic Disease; Clinical; Clinical Research; Clinical Trials; Colitis; Complex; Crohn' s disease; Data; Development; Diagnosis; Diet; Dietary Factors; Dietary Intervention; Dietary Supplementation; Disease Progression; Ellagi-Tannins; Ellagic Acid; Environmental Risk Factor; Epithelial; Fatty acid glycerol esters; Flare; Food; Future; Gnotobiotic; Gut Mucosa; Health; Human Microbiome; Immune; Immune response; Immune system; Immunity; In Vitro; Individual; Inflammatory; Inflammatory Bowel Diseases; Inflammatory Infiltrate; Intake; Interleukin-10; Intervention; Intestinal Mucosa; Intestines; Knock-out; Link; Mediating; Mesentery; Metabolic; Modeling; Molecular; Mucositis; Mucous Membrane; Mus; Nature; Nutrient; Pathogenesis; Pathway interactions; Patients; Pharmaceutical Preparations; Phenotype; Play; Pomegranate; Predisposition; Prevalence; Production; Protocols documentation; Rag1 Mouse; Relapse; Research; Resistance; Role; Shapes; Shotguns; Signal Transduction; Spontaneous colitis; Sucrose; Supplementation; Symptoms; T cell response; T-Lymphocyte; Therapeutic; Tight Junctions; Tissues; Transplantation; Ulcerative Colitis; Variant; Veterans; Wild Type Mouse; base; chemically induced colitis; cytokine; dietary; dietary supplements; disorder risk; efficacy evaluation; gut bacteria; gut inflammation; gut microbiome; gut microbiota; improved; individual response; inter-individual variation; intraperitoneal; metabolic phenotype; metabolomics; metagenomic sequencing; microbial; microbiome; microbiome alteration; microbiota; mouse model; murine colitis; novel; novel strategies; pathobiont; pathogen; preclinical study; prevent; response; symptomatic improvement; treatment response

The prevalence of inflammatory bowel disease (IBD), such as Crohn’s disease and ulcerative colitis, is increasing in Veterans. Diet plays an important role in shaping the intestinal microbiota and has emerged as an important and actionable modifier of IBD. In addition, metabolites derived from nutrients by gut bacteria have been shown to modify IBD progression. For example, pomegranate ellagitannins (ET) are broken down to ellagic acid (EA) and further converted to urolithins by gut bacteria. Both EA and UA were demonstrated to improve symptoms of IBD in chemically induced mouse models of colitis. We have recently shown that dietary pomegranate extract (PomX) supplementation reduced colitis markers and downregulated colitis associated inflammatory pathways in IL-10-/- mice. We also found that PomX and UA reduced pathobionts in wildtype mice. In addition, inter- individual variations in urolithin production, so called metabotypes (A: only produce UA, B: forms urolithin B (UB), or isoUA; 0: do not form any urolithins) have been well documented and associated with individual’s metabolic health. It is our hypothesis that dietary PomX and/or UA suppress IBD pathogenesis by changing the gut microbiome, strengthening the gut barrier integrity and inhibiting the differentiation of Th1 and Th17 intestinal T cells and pro-inflammatory cytokine. Therefore, we propose to investigate whether dietary supplementation with PomX and/or UA will prevent or alleviate colitis and to identify the mechanisms involved in PomX/UA-induced suppression of colonic inflammatory pathways. The IL-10-/- mouse model, which mirrors the multifactorial nature of IBD, is ideal to investigate the effect of dietary supplementation on colitis. To accomplish this, in aim 1 we will investigate the effect of PomX in suppressing spontaneous colitis development in IL-10-/- mice, which either produce UA (metabotye A) naturally, or lack bacteria forming UA (metabotype 0). We will also evaluate whether UA will enhance PomX effects in metabotype 0 mice. Last, we will interrogate the potential mechanism of action of PomX and/or UA supplementation on gut microbiome and host immunity by performing fecal shotgun metagenomics sequencing, fecal and blood metabolomics, colonic tissue snRNAseq, cytokine RTqPCR, tissue tight junction analysis and immune cell phenotyping in IL-10-/- mice receiving dietary PomX and/or UA. In aim 2, we will investigate whether PomX and/or UA induced changes in the gut microbiome are responsible for their effect on colitis suppression in IL-10-/- mice. Gnotobiotic IL10-/-mice will be colonized with naïve, or PomX, and/or UA treated gut microbiome from human donors (metabotype A and B), and fed a high fat/high sucrose (HFHS) diet for 12 weeks. We will investigate the role of donor microbiome metabotype to PomX treatment could play in colitis susceptibility. In aim 3, we will evaluate the effects of PomX and/or UA on the intestinal T cell- mediated immune inflammatory pathways associated with colitis. Rag1-/- mice will be pre-treated with dietary PomX and/or UA and then injected intraperitoneally with CD4 naïve cells isolated from wildtype mice. After the T cell transfer the mice will undergo the dietary intervention for 12 weeks and their course of colitis will be compared to mice fed the HFHS diet. To establish whether PomX and/or UA effects are mediated through the modulation of the microbiome or not, in vitro studies utilizing primary CD4+ cell lines will be used to evaluate PomX and/or UA direct effects on TCR activated CD4+ cell phenotype, and on the proliferative, signaling and cytokine responses. Our proposed research presents a novel approach using dietary PomX/UA to prevent or attenuate colitis by targeting the intestinal microenvironment, created by bacteria and their metabolites leading to selective interaction with the host immune system. Results from these preclinical studies will lay important groundwork for future clinical studies to explore the use of dietary PomX/UA supplements as alternative and adjuvant therapy to improve gut mucosa inflammation or alleviate relapse in IBD, thus reducing immune- suppressing medications to a minimum in patients with mild to moderately active IBD.

炎症性肠病(IBD)的患病率正在增加，如克罗恩病和溃疡性结肠炎 在退伍军人中。饮食在塑造肠道微生物区系中起着重要作用，并已成为一种重要的 和可操作的IBD修饰物。此外，肠道细菌从营养素中提取的代谢物也已被证明。 修改IBD进展情况。例如，石榴单宁(ET)被分解成鞣花酸(EA)。 并被肠道细菌进一步转化为尿石素。电针和尿酸均能改善患者的症状 化学诱导的小鼠结肠炎模型中的IBD。我们最近发现，饮食中的石榴提取物 (POMX)补充降低结肠炎标志物和下调结肠炎相关炎症途径 在IL-10-/-小鼠体内。我们还发现POMX和UA减少了野生型小鼠的致病力。此外，国际- 尿凝素产生的个体变异，即所谓的代谢型(A：仅产生UA，B：形成尿凝素B(UB)， 或isUA；0：不形成任何尿石素)已经被很好地记录并且与个体的新陈代谢有关 健康。我们的假设是饮食中的POMX和/或UA通过改变肠道来抑制IBD的发病 微生物组，增强肠道屏障完整性，抑制Th1/Th17肠道T细胞分化 细胞和促炎细胞因子。因此，我们建议调查饮食补充剂是否含有 Pomx和/或UA将预防或减轻结肠炎，并确定Pomx/UA诱导的机制 抑制结肠炎症通路。IL-10-/-小鼠模型，反映了多因素的性质 是研究饮食补充剂对结肠炎影响的理想方法。为了实现这一目标，在目标1中，我们将 探讨POMX对IL-10-/-小鼠自发性结肠炎的抑制作用 自然产生UA(代谢型A)，或缺乏形成UA(代谢型0)的细菌。我们还将评估是否 UA可增强代谢0型小鼠的POMX效应。最后，我们将询问潜在的作用机制 Pomx和/或UA补充剂对肠道微生物群和宿主免疫的影响 元基因组测序，粪便和血液代谢组学，结肠组织SnRNAseq，细胞因子RTqPCR，组织 接受饮食POMX和/或UA的IL-10-/-小鼠的紧密连接分析和免疫细胞表型。在目标2中， 我们将调查POMX和/或UA引起的肠道微生物群的变化是否与他们的 对IL-10-/-小鼠结肠炎的抑制作用。灵知生菌IL10-/-小鼠将被天真或庞氏定植， 和/或UA处理来自人类供体(代谢型A和B)的肠道微生物组，并喂饲高脂肪/高蔗糖 (HFHS)饮食12周。我们将研究供体微生物组代谢型在POMX治疗中的作用 发挥对结肠炎的易感性。在目标3中，我们将评估POMX和/或UA对肠道T细胞的影响。 与结肠炎相关的免疫炎症途径。Rag1-/-小鼠将接受膳食预处理 Pomx和/或UA，然后用从野生型小鼠分离的CD_4幼稚细胞腹腔注射。后 T细胞转移小鼠将接受为期12周的饮食干预，它们的结肠炎病程将 与喂食HFHS饮食的小鼠相比。以确定POMX和/或UA效应是否通过 微生物群的调节与否，将使用原代CD4+细胞系的体外研究来评估 POMX和/或UA直接影响TCR激活的CD4+细胞表型，以及对增殖、信号转导和 细胞因子反应。我们建议的研究提出了一种新的方法，使用膳食POMX/UA预防或 通过靶向肠道微环境来减轻结肠炎，这些微环境是由细菌及其代谢产物 与宿主免疫系统选择性地相互作用。这些临床前研究的结果将具有重要意义 为未来的临床研究探索膳食POMX/UA补充剂作为替代和 辅助治疗改善肠黏膜炎症或减轻IBD复发，从而降低免疫力。 将轻度至中度活动期IBD患者的用药减少到最低限度。