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.
Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease with no available cures. The intestine contains trillions of bacteria that influence many aspects of health and disease, and we have found that by affecting the bacteria in the gut in models of ALS, can alter disease progression. We will identify potential protective factors provided by the microbiota and investigate the immune mechanisms by which they act in hopes of identifying new therapeutic targets for treatment of ALS.
