Identification and characterization of gut microbial bioactive molecules that determine predisposition to autoimmune disease and atopy

确定自身免疫性疾病和特应性倾向的肠道微生物生物活性分子的鉴定和表征

• 批准号: 9238030
• 负责人: Jon Clardy
• 金额: $ 68.65万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2021-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9238030
• 关键词: Accelerometer; Address; Affect; Allergic Disease; Anti-Inflammatory Agents; Anti-inflammatory; Appearance; Archives; Autoimmune Diabetes; Autoimmune Diseases; Bacteria; Bioinformatics; Biological; Biological Assay; Biology; Biopsy; Brain; Cell Extracts; Cell Line; Cells; Cellular Stress; Chemicals; Coculture Techniques; Complex; Coupled; Crohn' s disease; Crystallization; Culture Media; Culture Techniques; Data; Data Set; Dendritic Cells; Development; Digestion; Disease; Disease Progression; Dose; Environment; Epithelial Cells; Feces; Finland; Fractionation; Gastrointestinal Diseases; Gene Cluster; Genetic; Genetic Predisposition to Disease; Hand; Heart; High Pressure Liquid Chromatography; Human; Hypersensitivity; Image; Immune System Diseases; Immune response; Immune system; Immunity; Incidence; Individual; Infant; Inflammation; Inflammatory; Inflammatory Bowel Diseases; Inflammatory Response; Infusion procedures; Insulin-Dependent Diabetes Mellitus; Intestines; Laboratories; Lead; Libraries; Link; Living Standards; Mass Spectrum Analysis; Measurement; Measures; Membrane; Metabolism; Methodology; Microbe; Modification; Molecular; Molecular Biology; Molecular Weight; Monitor; Mucous Membrane; Mucous body substance; Myeloid Cells; NIH Program Announcements; Neurotransmitters; Nuclear Magnetic Resonance; Organoids; Pathway interactions; Patients; Physiological; Play; Population; Porifera; Predisposition; Probiotics; Production; Public Health; Reporter; Role; Sampling; Signal Pathway; Signal Transduction; Signaling Molecule; Structure; System; TNF gene; Techniques; Therapeutic; Therapeutic Intervention; Ulcerative Colitis; Vitamins; Work; X ray diffraction analysis; X-Ray Diffraction; aqueous; atopy; base; body system; cell type; cohort; crystallinity; design; detector; gut microbiome; gut microbiota; immune activation; immunoregulation; insight; member; microbial; monolayer; novel; response; screening; small molecule; stem; therapeutic target; three dimensional structure; transcription factor; two-dimensional

ABSTRACT We know that the gut microbiome functions as an accessory organ system that plays critical roles in human digestion; the production of essential vitamins, neurotransmitters, and other small molecules; and in the regulation of the immune system. We also know that changes in the gut microbiota can lead to disease, especially inflammatory bowel disease and related inflammatory disorders. However, we know very little about the molecules and mechanisms that connect gut microbiota to these diseases. As the bacteria in the gut microbiota sense and respond to their environment with small molecules, it's likely that some of these molecules are key regulators of both pro-inflammatory and anti-inflammatory responses. This project seeks to identify these molecules and the mechanisms by which they operate. The project stems from a rich data set (DIABIMMUNE) that revealed a number of robust correlations between the appearance, or disappearance, of members of the gut microbiota and the development of dysregulated immune responses and disease in infants with genetic pre-disposition to type 1 diabetes. We will focus on ~40 strains that appear to play outsize roles in disease progression to discover what pro- or anti-inflammatory small molecule signals they produce. In Specific Aim 1 we will culture patient-derived strains of these bacteria under a variety of conditions, capture the small molecules produced, and measure their ability to regulate a pro-inflammatory signal (TNFalpha) and an anti- inflammatory signal (IL-10). The most potent and selective regulators will advance to Specific Aim 2 in which bioassay-guided fractionation will guide the isolation of active molecules, which will then be structurally characterized with spectroscopic (both nuclear magnetic resonance and mass spectrometry) and imaging (X-ray diffraction) techniques. With known molecules linked to confirmed biological activity in hand, we will prioritize hits based on their presence in the DIABIMMUNE stool samples, which we have archived. Only bioactive molecules present in human samples will be analyzed in specific Aim 3 to determine mechanism. Because the signaling pathways through which these molecules exert their biological effects are complex, we will employ a specially designed and newly implemented assay that simultaneously measures more than 50 transcription factors that regulate inflammation, immunity, metabolism and cell stress. By profiling transcription factor activity and comparing to known compounds, we will be able to classify molecules based on their activity profiles. We will further characterize their mechanism of action using a variety of functional assays on both patient derived myeloid cells as well as colonic and ileal mucosa cell lines that develop into four distinct cell types that form three-dimensional structures including crypts covered by a secreted mucus layer. At the end of the pipeline formed by these three specific aims, small molecules that regulate immune responses along annotated pathways will be known, and this identification of both molecule and mechanism will provide insights into both basic biology and potential therapeutic interventions.

抽象的 我们知道，肠道微生物组充当辅助器官系统，在人类中起着关键作用 消化;必需维生素，神经递质和其他小分子的产生；在 免疫系统的调节。我们还知道，肠道菌群的变化会导致疾病， 特别是炎症性肠病和相关炎症性疾病。但是，我们对 将肠道菌群与这些疾病联系起来的分子和机制。作为肠道中的细菌 微生物群感并用小分子对环境做出反应，其中一些分子很可能 是促炎和抗炎反应的关键调节剂。该项目旨在确定 这些分子及其运行的机制。该项目源于丰富的数据集 （Diabmmune）揭示了外观或消失的许多牢固相关性 肠道菌群的成员以及婴儿免疫反应和疾病失调的发展 与1型糖尿病的遗传预求。我们将专注于约40条似乎扮演角色的菌株 疾病进展，发现它们产生的促或抗炎小分子信号。具体 目的1我们将在各种疾病下培养这些细菌的患者衍生菌株，捕获小的 产生的分子并测量其调节促炎信号（TNFALPHA）的能力和抗 炎症信号（IL-10）。最有力，最有选择的调节器将晋升为特定目标2 生物测定引导的分级将指导活性分子的分离，然后在结构上是 以光谱镜（核磁共振和质谱法）和成像（X射线）的特征 衍射技术。随着已知的分子与确认的生物学活性有关，我们将优先考虑 基于它们在我们存档的糖尿病粪便样品中的存在。只有生物活性 人类样品中存在的分子将在特定目标3中进行分析，以确定机制。因为 这些分子发挥其生物学作用很复杂的信号通路，我们将采用一个 专门设计和新实施的测定法，同时测量50多个转录 调节炎症，免疫力，代谢和细胞应激的因素。通过分析转录因子活动 与已知化合物相比，我们将能够根据它们的活性曲线对分子进行分类。我们 将进一步表征他们的作用机理，并在两个患者衍生的情况下使用各种功能测定 髓样细胞以及结肠和回肠粘膜细胞系，它们形成四种不同的细胞类型 三维结构，包括被分泌的粘液层覆盖的隐窝。在管道结束时 由这三个特定目的形成，小分子沿着注释 途径将是已知的，分子和机制的这种识别将为两者提供见解 基本生物学和潜在的治疗干预措施。