TRANQUIL attenuates autoimmune demyelination and suppresses neuroinflammatory signaling in experimental autoimmune encephalomyelitis.

Neuroinflammation driven by autoreactive T cells and activated glial populations underlies the pathogenesis of multiple sclerosis (MS), yet current immunomodulatory therapies rarely restore durable immune tolerance and often cause broad immunosuppression. We previously engineered an attenuated Brucella melitensis ΔvjbR strain expressing tnaA, enabling constitutive production of the tryptophan metabolite indole, a modulator of antigen-presenting cell (APC) activation and T-cell differentiation. Here, we tested whether this strain (termed TRANQUIL - Technology for Reprogramming Autoimmunity And Quieting Inflammation, TRQ) ameliorates neuroinflammation in experimental autoimmune encephalomyelitis (EAE). Female C57BL/6 mice with EAE received a single intravenous dose of TRQ, and PBS-treated mice served as comparator controls. TRQ treatment was associated with reduced clinical disease severity during the observation period. Histological analysis of spinal cord sections showed moderately reduced inflammatory infiltration and partial preservation of myelin architecture compared with PBS-treated controls. Quantitative immunohistochemical analysis showed reduced Iba1 and GFAP immunoreactivity in treated mice. Bulk RNA sequencing of spinal cords demonstrated broad downregulation of inflammatory and immune-related pathways, including NF-κB, TNF, NOD-like receptor, cytokine-cytokine receptor interaction, and T-cell differentiation pathways. Biodistribution studies showed transient systemic distribution and rapid clearance, with no detectable bacteria in brain or spinal cord. Together, these findings indicate that TRQ administration is associated with partial attenuation of EAE severity, reduced CNS inflammatory pathology, and broad suppression of inflammatory transcriptional signatures. Additional studies incorporating appropriate bacterial controls and mechanistic comparisons will be required to determine the contribution of engineered indole production relative to broader immune effects elicited by systemic bacterial exposure.