Background/Purpose: Intestinal microbial dysbiosis, intestinal inflammation, and Th17 immunity are all linked to the pathophysiology of ankylosing spondylitis (AS); however, the mechanisms linking them remain unknown. Using three unbiased approaches, we hypothesized that microbial dysbiosis in AS results in altered bacterial metabolites that expand IL-17-producing immune cells.

Methods: Healthy controls (HC, N=23) and patients with AS (N=24) were recruited while undergoing standard-of-care colonoscopy. Thirty distal colon biopsies and rectal swabs were obtained from each subject. Four biopsies were submitted for metabolomics screening via LC-MS while the remaining were homogenized and analyzed by single cell RNA sequencing (scRNA-seq, 2 subjects per group) and flow cytometry (remaining) to validate findings. Bacterial DNA was extracted from the rectal swabs, underwent shotgun sequencing, and analyzed using the HumanN2 software suite.

Results: ScRNA-seq identified a unique population in AS versus HC that transcriptionally segregated with T cells but lacked traditional T cell markers. Flow cytometry validated a significant expansion of type 3 innate lymphoid cells (ILC3s) in the intestinal tissue from patients with AS. By LC-MS, multiple metabolites within the tryptophan pathway were found to be significantly increased in AS vs HC, including indole-3-acetate (relative tissue concentration 153,583±156,338 vs 36,124±29,764, p< 0.0003) and indole-3-acetaldehyde (relative tissue concentration 2,615,095±1,890,967 vs 1,068,847±1,158,350, p< 0.0005), which are bacteria-specific products of tryptophan metabolism. Metagenomic analysis of the bacterial populations in AS identified altered metabolic pathways involving amino acid synthesis/degradation. In particular, there was a predilection towards tryptophan synthesis in HC while other tryptophan metabolism pathways were altered in AS.

Conclusion: In this study we identified altered tryptophan metabolism with a corresponding increase in ILC3s in intestinal tissue from patients with AS, which may link intestinal pathology to Th17 immunity in AS. Future studies will focus on the mechanistic pathways between tryptophan metabolism and ILC3 development.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/metabolic-regulation-of-type-3-innate-lymphoid-cells-by-intestinal-bacteria-derived-indoles-in-ankylosing-spondylitis/