Sunday, June 1, 2025
6/1/2025
AhR suppression of T-bet+ B cells in SLE - Project Summary: This proposal will answer two key questions. 1. Does IL-4 act through the aryl hydrocarbon receptor (AhR) pathway to inhibit the development of double negative 2 (DN2) B cells? 2. How does IL-4 induce endogenous ligands of AhR to suppress T-bet+ B cells? We recently showed that in SLE, down-regulation of IL- 4R and defective maintenance of resting naïve (rNAV) B cells were associated with increased development of Tbet+CD11c+ activated naïve (aNAV) B cells and DN2 B cells. Co-culture of SLE B cells with IL-4 promoted the maintenance of rNAV B cells and blocked both type I and type II interferon (IFN)-promoted development of aNAV and DN2 B cells. Single-cell transcriptomics of lupus mouse B cell stimulated in vivo with IL-4 revealed up- regulation of IL-4-induced1 (IL4i1). As an L-amino acid oxidase, IL4i1 can catalyze the synthesis of indole-pyruvic acid (IPyA) derivative endogenous AhR ligands leading to upregulation of AhR response genes. Metabolome analysis revealed that IL-4 induced AhR agonistic metabolites in B cells, including kynurenine (Kyn) and indole- 3 metabolites. In the absence of IL-4, AhR agonists 6-Formylindolo[3,2-b]carbazole (FICZ) and Kyn significantly suppressed IFNβ and TLR7-induced T-bet+CD11c+ IgD− B-cell development. We hypothesize that IL-4 activates AhR through a combined effect of IL4i1 and IDO1 to promote the formation of endogenous AhR ligands in B cells. In Aim 1, we will determine if AhR is required to inhibit T-bet+ B cell development in mice and what pathways and metabolites are involved. This will be tested using conditional AhR-deficient mice under chronic TLR7 stimulation conditions (Aim 1a). The pathway for the production of AhR ligands, including indole-3 metabolites and Kyn, will be determined using Il4i1−/− mice and Ido1−/− mice (Aim 1b). The ability of AhR agonistic metabolites to regulate T-bet+ B cell development in vivo will also be tested (Aim 1c). In Aim 2, we will use B cells derived from SLE patients to determine if AhR is required for IL-4 to suppress DN2 B cell development (Aim 2a). We will further determine if modulation of IL4I1, IDO1, and their downstream metabolites can alter the development of DN2 B cells (Aim 2b). Significance. A primary focus in SLE research has been studies of elevated pro-inflammatory factors that drive pathogenic autoimmune responses. We propose that therapies designed to promote long-term homeostasis in otherwise abnormally stimulated immune cells could be more efficacious and less toxic. Identifying small molecular metabolites that act directly in B cells to induce homeostasis may lead to the development of orally active druggable targets that are efficacious in treating SLE. Innovation. The proposed project is novel as B-cell endogenous metabolic ligands that can maintain B-cell homeostasis and suppress T-bet+ B-cell development are unknown. Team and Environment. Drs. Mountz, Rubio, and Hsu have a history of collaboration to study cytokine-mediated B cell single cell transcriptomics and pathogenesis in SLE. Dr. Barnes, Director of the UAB Targeted Metabolomics and Proteomics Laboratory (TMPL), will provide expertise in metabolomics analysis.
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