Role of the Intestinal Microbiota in ALS

项目总结 肌萎缩侧索硬化症是一种影响运动神经元的神经退行性疾病，治疗选择有限，中位生存期 3-5年。我们提交了一份经过重大修改的提案，以回应审查员的意见，并 提供了新的初步数据。我们的假设是肠道微生物及其代谢物在 通过调节外周和中枢神经系统免疫及影响ALS疾病在ALS中的重要作用 小路。Blacher最近在《自然》杂志上发表的一篇论文证实了我们的假设，该论文显示了 肌萎缩侧索硬化症的肠道微生物群和代谢产物。在新的初步数据中，我们显示：1)抗生素会恶化存活率 同时下调小胶质细胞稳态基因，同时上调炎症基因。2)更改 68名ALS患者与61名健康对照的微生物组(迄今为止最大的微生物组研究)，包括减少 在丁酸产生菌中，E.rectale和R.enterinalis在控制ALS临床时是健壮的 混血儿。3)接种这些细菌或阿克曼草能逆转SOD1病相关的转录 脊髓的变化；包括FUS、Oxr1和SMN1，以及蛋白质降解(Ubqln1)。4)调入 SOD1微生物群到WT小鼠调节与RNA加工(FUS)相关的ALS中的小胶质细胞通路， 蛋白质降解(HSPa1b和USP2)和溶酶体运输(CD68和Lyz2)。我们相信有 令人信服的证据支持对肌萎缩侧索硬化症微生物群的研究。我们将致力于实现这些目标： 目的1.在SOD1和TDP-43模型中，哪些微生物成分与保护相关？ 我们将用特定的低剂量抗生素耗尽微生物区系，口服阿克曼草组分和 一种独特的微型RNA，可识别与疾病保护相关的脑、血清和粪便代谢物。 目的2.哪些人类微生物区系在疾病发病机制中起作用？ 目前尚不清楚ALS微生物区系是否也可以驱动疾病的发病机制。我们将把微生物区系从 ALS患者采用SOD1和TDP-43模型，测量运动功能和生存时间。我们会 确定与保护或恶化疾病相关的微生物种群、功能和代谢物 疾病。我们还将在一个新招募的队列中确认和扩大我们在ALS微生物群中的发现。 目的3.研究肠道微生物区系调控疾病的免疫机制 ALS动物模型的研究进展。我们将对小鼠的小胶质细胞、单核细胞和T细胞进行分选 单独的抗生素，与ALS微生物群或特定的细菌菌株共同定植，并表征转录 由RNAseq签名。利用WT小鼠研究微生物区系如何影响小胶质细胞，我们将转移 确定与肌萎缩侧索硬化症相关的特定微生物，并筛选参与肌萎缩侧索硬化症发病机制的基因的变化。

PROJECT SUMMARY ALS is a neurodegenerative disease affecting motor neurons with limited treatment options and a median survival of 3-5 years. We submit a significantly revised proposal that addresses the comments of the reviewers and provides new preliminary data. Our hypothesis is that the intestinal microbiome and its metabolites play an important role in in ALS by modulating peripheral and CNS immunity and by affecting ALS disease pathways. Our hypothesis is strengthened by a recent paper in Nature by Blacher showing an important role of the gut microbiome and metabolites in ALS. In new preliminary data we show: 1) Antibiotics that worsen survival also downregulate microglia homeostatic genes while upregulating inflammatory genes. 2) Changes in the microbiome in 68 ALS patients vs. 61 healthy controls (largest microbiome study to date), including a decrease in butyrate producing bacteria E. rectale and R. intestinalis, are robust when controlled for ALS clinical confounders. 3) Administering these bacteria or Akkermansia reverses SOD1-disease associated transcriptional changes in the spinal cord; including Fus, Oxr1, and Smn1, and protein degradation (Ubqln1). 4) Transferring SOD1 microbiota to WT mice modulates microglia pathways involved in ALS related to RNA processing (Fus), protein degradation (HSPa1b and USP2) and lysosomal transport (CD68 and Lyz2). We believe there is compelling evidence to support the investigation of the microbiome in ALS. We will address these aims: AIM 1. Which microbial components are associated with protection in SOD1 and TDP-43 models? We will deplete the microbiota with specific low-dose antibiotics, orally administer Akkermansia components and a unique micro-RNA and identify brain, serum, and stool metabolites associated with disease protection. AIM 2. Which human microbiota components contribute to disease pathogenesis? It is unknown whether the ALS microbiota can also drive disease pathogenesis. We will transfer microbiota from patients with ALS to the SOD1 and TDP-43 models and measure motor function and survival time. We will identify microbial populations, functions, and metabolites that are associated with protection or worsening of disease. We will also confirm and expand our findings in the ALS microbiome in a newly recruited cohort. AIM 3. Investigate the immune mechanisms by which the gut microbiota modulates disease progression in ALS animal models. We will sort microglia, monocytes, and T cells from mice treated with individual antibiotics, colonized with ALS microbiota, or specific bacterial strains and characterize transcriptional signatures by RNAseq. Utilizing WT mice to investigate how the microbiota affect microglia, we will transfer specific microbes identified associated with ALS and screen for changes in genes involved in ALS pathogenesis.