Understanding the role of aromatic amino acid derived metabolic toxins in neuroinflammation

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Project Summary Multiple sclerosis (MS) is the most common neurodegenerative disease in young adults. Changes in the serum metabolome have been reported in MS and our preliminary data demonstrate that they are also associated with MS disease severity. We specifically show that aromatic amino acid (AAA)-derived metabolic toxins, produced by the gut microbiota, are associated with greater disease severity. Our long-term goals are to improve the understanding of the role of circulating metabolites in the pathophysiology of MS and discover novel therapeutic interventions based on those findings. The objectives of this R21 application are to determine whether AAA- derived metabotoxins have direct pro-inflammatory effects on immune and glial cells in vitro and also to determine whether treatment with these metabolites worsens neuroinflammation in a mouse model of MS. The rationale for this project, supported by preliminary data, is that people with MS have higher levels of these AAA- derived metabotoxins in their blood compared to healthy controls and the levels of these metabotoxins are correlated with disease severity measured clinically and by imaging measures. The proposed research study will pursue two specific aims: 1) to determine whether AAA-derived metabotoxins have pro-inflammatory effects on immune and glial cells in vitro; 2) To determine whether AAA-derived metabotoxins worsen the severity of neuroinflammation in an animal model of MS. For the first aim, we will test the effects of a variety of concentrations of four AAA-derived metabotoxins on pro-inflammatory polarization of adaptive (T and B cells) and innate (macrophages) immune cells, as well as glial cells (astrocytes and microglia). We will compare effects of various metabolites to vehicle and identify those that have pro-inflammatory effects (especially those with a dose-response relationship) without affecting cell viability. For the second aim, we will test the effects of supplementation with these AAA-derived metabotoxins on the severity of neuroinflammation in an animal model of MS – experimental autoimmune encephalomyelitis (EAE). We will utilize outcomes of inflammation (inflammatory cell infiltration and demyelination) and neurodegeneration (retinal ganglion cell counts in retinal flat mounts) in EAE mice. This will help provide complementary data to those generated in Aim 1. This project is innovative in that it proposes to test a novel hypothesis that metabolites in the circulation that are associated with MS disease severity may have direct effects on immune and glial cell function and may also affect neuroinflammation in an animal model of MS. The proposed research is significant because it can provide novel understanding of the mechanisms underlying MS disease pathogenesis and identifies new strategies for therapeutic interventions (such as dietary changes, probiotics, antibiotics and metabolite supplementation) for this common disabling neurological disorder.