Multi ‘Omics Analysis of Intestinal Tissue in Ankylosing Spondylitis Identifies Alterations in the Tryptophan Metabolism Pathway

Abstract

Intestinal microbial dysbiosis, intestinal inflammation, and Th17 immunity are all linked to the pathophysiology of spondyloarthritis (SpA); however, the mechanisms linking them remain unknown. One potential hypothesis suggests that the dysbiotic gut microbiome as a whole produces metabolites that influence human immune cells. To identify potential disease-relevant, microbiome-produced metabolites, we performed metabolomics screening and shotgun metagenomics on paired colon biopsies and fecal samples, respectively, from subjects with axial SpA (axSpA, N=21), Crohn’s disease (CD, N=27), and Crohn’s-axSpA overlap (CD-axSpA, N=12), as well as controls (HC, N=24). Using LC-MS based metabolomics of 4 non-inflamed pinch biopsies of the distal colon from subjects, we identified significant alterations in tryptophan pathway metabolites, including an expansion of indole-3-acetate (IAA) in axSpA and CD-axSpA compared to HC and CD and indole-3-acetaldehyde (I3Ald) in axSpA and CD-axSpA but not CD compared to HC, suggesting possible specificity to the development of axSpA. We then performed shotgun metagenomics of fecal samples to characterize gut microbial dysbiosis across these disease states. In spite of no significant differences in alpha-diversity among the 4 groups, our results confirmed differences in gene abundances of numerous enzymes involved in tryptophan metabolism. Specifically, gene abundance of indolepyruvate decarboxylase, which generates IAA and I3Ald, was significantly elevated in individuals with axSpA while gene abundances in HC demonstrated a propensity towards tryptophan synthesis. Such genetic changes were not observed in CD, again suggesting disease specificity for axSpA. Given the emerging role of tryptophan and its metabolites in immune function, altogether these data indicate that tryptophan metabolism into I3Ald and then IAA is one mechanism by which the gut microbiome potentially influences the development of axSpA.

Axial spondyloarthritis patients have altered mucosal IgA response to oral and fecal microbiota

“…Notably, IgA-SEQ revealed many microbes that were not differentially abundant using the 16S sequencing alone, e.g., Akkermansia, and members of family Ruminococcaceae and Lachnospiraceae. These microbes have been shown to be associated with clinical studies in SpA and IBD (12,31,32) as well as in experimental models of SpA (22,33,34). Interestingly, Akkermansia has been implicated as a pathogenic microbe in SpA and IBD (12,31,32); and a protective microbe in metabolic diseases and obesity studies (35,36).…”

“…These microbes have been shown to be associated with clinical studies in SpA and IBD (12,31,32) as well as in experimental models of SpA (22,33,34). Interestingly, Akkermansia has been implicated as a pathogenic microbe in SpA and IBD (12,31,32); and a protective microbe in metabolic diseases and obesity studies (35,36). We also observed other IgA coated microbes in the fecal (e.g., Lachnospiraceae, Ruminococcaceae) and salivary (e.g., Prevotellaceae, Actinobacillus) samples from AxSpA patients, which have been shown to be associated with various spondyloarthropathies (12,14).…”

Intestinal Microbial Metabolites in Ankylosing Spondylitis

“…Nevertheless, it is worth noticing that two main amino acid alterations were confirmed in different “omics” studies (reported in Table 1 ): glutamine and tryptophan (Trp). In particular, Gao et al found a decreased level of Trp in plasma of AxSpA patients compared to healthy controls [ 22 ], suggesting an increased metabolization of the Trp by the host cells, while Berlinberg et al, using intestinal tissue samples and matched fecal samples metagenomics studies, detected significant alterations in Trp pathway metabolites sustained by bacteria metabolism [ 26 ].…”

“…Moreover, 4–6% of total Trp can be directly metabolized by gut bacteria, through the action of the enzyme tryptophanase, into tryptamine and indolic byproducts [ 26 , 105 ]. Some of these indole metabolites, such as IPA (indole propionic acid) and IAA (indole acetic acid), act as a ligand for the AhR and pregnane X receptor (PXR), regulating intestinal barrier function [ 107 , 119 , 120 ].…”

“…Consistently, IAA levels were reduced in Card9 −/− mice and the transfer of Card9 −/− intestinal microbiota to germ-free wild type mice was sufficient to impair IL-22 activation and to increase the sensitivity to colitis [ 119 ]. Moreover, a recent study associating intestinal samples metabolomics with matched fecal samples metagenomic profiling, demonstrated how microbiota from ax-SpA patients switched from Trp synthesis to Trp metabolism, compared to both healthy controls and Crohn’s disease controls, suggesting that this alteration is specific to ax-SpA regardless of the presence of bowel inflammatory diseases [ 26 ].…”