Microbiome and intestinal barrier in ALS therapy

本申请的目的是研究肠道微生物稳态在ALS中的重要作用， 同时研究一种新的方法，可以通过恢复宿主-微生物关系来治疗ALS 并与FDA批准的药物结合。肌萎缩侧索硬化症是一种致命的神经肌肉疾病 以运动神经元的进行性死亡和肌肉萎缩为特征。大多数患者在5年内死亡 在疾病发作后。目前，阿曲唑和Radicava治疗仅延长了少数患者的寿命。 个月因此，迫切需要开发用于ALS的新疗法并改善患者的生活质量。 ALS患者的质量。新出现的证据表明，微生物组和肠道内稳态 在神经系统疾病中起着重要作用，如阿尔茨海默病和帕金森病。然而，在这方面， 对ALS患者的肠道微生物组知之甚少。我们的实验室是第一个发现 在ALS小鼠模型G93 A中肠内稳态和疾病进展之间的关系。我们的研究在 人类ALS进一步揭示了生态失调和肠道炎症。ALS小鼠的肠道受损 结构和增加肠道通透性（肠漏）。值得注意的是，恢复肠道内稳态， 用细菌产物丁酸盐喂养ALS模型小鼠显著延迟了疾病的发作， 延长ALS小鼠的寿命。我们假设，针对肠道微生物组并结合FDA 批准的药物，以改善肠道屏障功能和恢复微生物组，从而减缓疾病 ALS的进展。这些研究旨在严格检验假设，并解决两个主要问题。 目标.目的1是确定导致ALS微生态失调和屏障功能障碍的机制。 我们将定义异常上皮连接结构和生态失调的机制（例如丁酸盐损失）。 在ALS小鼠中产生细菌）。我们将研究丁酸盐和丁酸盐产生细菌的作用，使用 新的分子工具、肠样培养物和转基因动物模型（SOD 1G 93小鼠和一种新的肠道特异性 S 0 D1 G93模型的过表达）。我们将确定细菌产物的益处的潜在机制， 丁酸对肠通透性和神经保护的影响。目标2是进行原理验证研究， 恢复肠道微生物组和肠屏障，以保护ALS的神经肌肉功能。ALS小鼠， ALS的不同阶段将用有益细菌（促进丁酸盐产生的益生菌）治疗，或 与FDA药物（阿舒唑或Radicava）联合使用。ALS小鼠将进行粪便微生物群移植（FMT） 使用健康的野生型小鼠作为供体。肠上皮细胞的生理和分子生物学评估 完整性、微生物组和神经肌肉性能将用于评价治疗效果。 将使用组织学和生物化学评价来确定肠上皮细胞水平的变化。 和肌萎缩侧索硬化症不同阶段的神经肌肉功能。我们将优化恢复肠道的功效 微生物组与FDA药物联合使用不同ALS小鼠模型减缓ALS进展 （SOD 1G 93 A和TDP 43）。退伍军人被诊断患有ALS的可能性是普通人群的两倍。 退伍军人，无论服役的分支，服役的时代，或者是否服役 在和平或战争时期，他们死于ALS的风险比没有在军队服役的人更大。 我们的研究是有意义的，因为ALS在VA人群中的健康负担和 微生物组对健康和疾病中的神经肌肉功能的影响，特别是在退伍军人护理中。创新点 该项目在于：（a）ALS的治疗潜力，（B）发现早期变化的概念框架 和生态失调以及ALS发作前的肠道屏障作用，以及（c）最先进的实验模型 这使我们能够了解益生菌和/或细菌的有益作用的新机制， 改善运动神经元功能的产品。它对开发新的 治疗策略，以对抗这种毁灭性的疾病，改善退伍军人的健康。

The goal of this application is to investigate the important role of intestinal microbial homeostasis in ALS, meanwhile investigating a novel approach that can potentially treat ALS by restoring host-microbe relationships and combining with FDA approved drugs. Amyotrophic lateral sclerosis (ALS) is a fatal neuromuscular disease characterized by progressive death of motor neurons and muscle atrophy. Most patients die within 5 years after the disease onset. Currently, treatment with Riluzole and Radicava only extends patient life span for a few months. Therefore, there are significant needs to develop novel treatments for ALS and improving the life quality of ALS patients. Emerging evidence has demonstrated that microbiome and intestinal homeostasis plays essential roles in neurological diseases, such as Alzheimer's disease and Parkinson's disease. However, little is known about the intestinal microbiome in patients with ALS. Our lab is the first to discover the link between intestinal homeostasis and the disease progression in an ALS mouse model G93A. Our study in human ALS further reveals the dysbiosis and intestinal inflammation. ALS mice had damaged intestinal structure and increased intestinal permeability (leaky gut). Remarkably, restoring the intestinal homeostasis by feeding the ALS model mice with a bacterial product butyrate significantly delayed the disease onset and prolonged the life span of ALS mice. We hypothesize that targeting gut microbiome and combining with FDA approved drugs to improve the intestinal barrier function and restore microbiome, thus slowing disease progression of ALS. The studies are designed to rigorously examine the hypotheses and address two main objectives. Aim 1 is to determine the mechanisms that contribute to dysbiosis and barrier dysfunction in ALS. We will define the mechanism for abnormal epithelial junction structure and dysbiosis (e.g. loss of butyrate- producing bacteria) in ALS mice. We will investigate roles of butyrate and butyrate-producing bacteria, using novel molecular tools, enteroid cultures, and transgenic animal models (SOD1G93 mice and a novel gut-specific overexpression of SOD1G93 model). We will determine mechanisms underlying the benefits of bacterial product butyrate on intestinal permeability and neuroprotection. Aim 2 is to conduct proof-of-principle studies with restoring gut microbiome and intestinal barrier for preserving neuromuscular function in ALS. ALS mice at different stages of ALS will be treated with beneficial bacteria (probiotics that promote butyrate production) or combining with FDA drugs (Riluzole or Radicava). ALS mice will have fecal microbiota transplantation (FMT) using health wild-type mice as donors. Physiological and molecular biological assessments of intestinal integrity, microbiome, and neuromuscular performance will be used to evaluate the therapeutic effects. Histopathology and biochemical evaluations will be used to determine changes at the cellular level of intestinal and neuromuscular function at different stages of ALS. We will optimize the efficacy of restoring intestinal microbiome combining with FDA drugs in slowing ALS progression using different ALS mouse models (SOD1G93A and TDP43). Veterans are twice as likely to be diagnosed with ALS as the general population. Military veterans, regardless of the branch of service, the era in which they served, or whether they served during a time of peace or war, are at a greater risk of dying from ALS than if they had not served in the military. Our study is significant because of the health burden of ALS in VA population and the novel role of the microbiome on neuromuscular function in health and disease, especially in Veterans' care. Innovation of this project lies in its: (a) therapeutic potential for ALS, (b) conceptual frame-work to discover early changes and dysbiosis and the gut barrier effects before onset of ALS, and (c) state-of–the-art experimental models that allow us to understand novel mechanisms underlying the beneficial effect of probiotics and/or bacterial products that improve the motor neuron function. It has significant translational implications for developing new therapeutic strategies for combating this devastating disease and improving the health of veterans.