Dark GPCR signaling underlying the Microbiome-Gut-Brain Axis for Alzheimer's Disease and Related Dementia

项目摘要 累积的证据表明微生物组-肠道-大脑轴在阿尔茨海默病中起着至关重要的作用 (AD)并支持微生物群靶向疗法作为AD和AD相关痴呆治疗的潜力 （ADRD）.然而，微生物组-肠道-脑轴的精确机制和可操作肠道的识别 AD/ADRD发病机制、疾病进展和修饰的微生物生物标志物提示 替补演员化学基因组学技术的最新进展表明，G蛋白偶联 受体（GPCR，人类基因组中最大的药物靶家族，由NIH资助的 阐明可药用基因组计划）介导了微生物组-肠道-大脑轴的大部分，特别是对于 肠道微生物群衍生的代谢物，例如中链脂肪酸（MCFA）。我们的初步实验 揭示了肠道微生物群MCFA代谢物之间的强烈显著关联（例如，5-苯基戊酸）和 暗GPCR信号传导（例如，GPR 84）在AD中的应用以及AD患者诱导的多能 干细胞（iPSC）模型。此外，我们确定了靶向肠道微生物代谢途径， 无菌小鼠的认知行为。我们认为，结合AD患者诱导的iPSC，脑类器官， 和无菌小鼠模型，沿着血浆和海马肠道微生物的多模式分析， 代谢组学数据，将能够提高对精确的保护机制的机械理解， AD/ADRD中的微生物组-肠道-脑轴。我们的中心统一假设是， 驱动器（例如，肠道微生物代谢物）和可药用GPCR信号网络的微生物- 肠-脑轴将引出AD潜在的预防和治疗策略。目标1将测试暗GPR 84（a 假定的小胶质细胞基因）及其信号传导激活，其是MCFA的微生物组-肠-脑轴的基础， 通过评估AD相关的认知和病理变化， 表型和机制。我们将评估不同的肠道微生物群落，非靶向和靶向 GPR 84-/-、5xFAD和杂交（5xFAD; GPR 84-/-）中血浆和海马体的肠道代谢组学分析 在FMT前和FMT后期间的无菌小鼠。Aim 2将筛选、测试和验证暗GPCR和信号 使用AD患者来源的iPSC系的肠道微生物群来源的MCFA代谢物的网络扰动 与脑类器官模型结合。具体来说，我们将评估肠道微生物的物理结合， 使用互补钙流、cAMP glosensor和β-arrestin Tango的代谢物-GPCR相互作用组 测定。目标3将对肠道微生物代谢物生物标志物的发现进行监督分析， 使用靶向和非靶向患者血浆的临床相关AD病理学特征的预测建模 肠道微生物代谢组学总之，我们的多学科方法包括无菌小鼠、AD患者- 衍生的iPSC和脑类器官模型，沿着人血浆肠道微生物代谢组学，将确定 潜在的针对微生物组的预防和治疗方法将在AD患者中直接进行测试。

PROJECT SUMMARY Cumulative evidence indicates the Microbiome-Gut-Brain axis plays a crucial role in Alzheimer’s disease (AD) and supports the potential of microbiome-targeted therapies as treatments for AD and AD-related dementia (ADRD). However, the precise mechanism of the Microbiome-Gut-Brain axis and the identity of actionable gut microbial biomarkers underlying AD/ADRD pathogenesis, disease progression, and modification remind understudied. Recent advances in chemogenomic technologies have demonstrated that G-protein-coupled receptors (GPCRs, the largest druggable target family in the human genome, as defined by the NIH-funded Illuminating the Druggable Genome Program) mediate much of the Microbiome-Gut-Brain axis, especially for gut microbiota-derived metabolites such as medium-chain fatty acids (MCFAs). Our preliminary experiments reveal strong significant associations between gut-microbiota MCFA metabolites (e.g., 5-phenylvaleric acid) and dark GPCR signaling (e.g., GPR84) in AD using multi-omics approaches and an AD patient-induced pluripotent stem cells (iPSC) model. Furthermore, we identified targeting gut microbial metabolite pathways improve cognitive behaviors in germ-free mice. We posit that combining AD patient-induced iPSC, cerebral organoids, and germ-free mouse models, along with multimodal analyses of plasma and hippocampus gut microbial metabolomics data, will enable improved mechanistic understanding of precise protective mechanisms of the Microbiome-Gut-Brain axis in AD/ADRD. Our central unifying hypothesis is that identifying likely molecular drivers (e.g., gut microbial metabolites) and druggable GPCR signaling networks underlying the Microbiome- Gut-Brain axis will elicit potential prevention and treatment strategies for AD. Aim 1 will test dark GPR84 (a putative microglial gene) and its signaling activation underlying the Microbiome-Gut-Brain axis of MCFAs via fecal microbiota transplantation (FMT) in germ-free mice by assessing AD-related cognitive and pathological phenotypes and mechanisms. We will evaluate differential gut microbial communities, untargeted and targeted gut metabolomics analyses of plasma and hippocampus in GPR84-/-, 5xFAD, and cross (5xFAD;GPR84-/-) germ-free mice during pre-FMT and post-FMT. Aim 2 will screen, test and validate dark GPCRs and signaling network perturbation by gut microbiota-derived MCFA metabolites using AD patient-derived iPSC lines in conjunction with cerebral organoid models. Specifically, we will evaluate physical binding of the gut microbial metabolite-GPCR interactome using complementary Calcium flux, cAMP glosensor, and β-arrestin Tango assays. Aim 3 will conduct supervised analyses of gut microbial metabolite biomarker discovery for, and prediction modeling of, clinically relevant AD pathological features using patient plasma targeted and untargeted gut microbial metabolomics. In summary, our multidisciplinary approach comprising germ-free mice, AD patient- derived iPSC, and cerebral organoid models, along with human plasma gut microbial metabolomics, will identify potential microbiome-targeted prevention and treatment approaches to be directly tested in people with AD.