Background/Purpose: The strong clinical and genetic associations between axial spondyloarthritis (axSpA) and inflammatory bowel disease (IBD) underscore the pathogenic role of the gut-joint axis. We previously identified a subpopulation of pathogenic mature CD8+ T cells in the axSpA synovial fluid (SF), expressing CD103+ and CD49a+ integrins (InEx cells). The expression of integrins on InEx cells implicates their role in aberrant migration between the gut-joint axis. Whether InEx cells recognize an arthritogenic peptide remains unclear. We hypothesize that InEx cells may incite and perpetuate chronic inflammation in axSpA. To this end, we characterized their T-cell receptor (TCR) repertoire and gene expression profile.

Methods: Patients fulfilling the ASAS classification for axSpA were recruited (n=4). Active joint effusion from these patients is reflective of active joint inflammation which coincided with high disease activity (BASDAI >4). We isolated peripheral blood mononuclear cells (PBMCs) and synovial fluid mononuclear cells (SFMCs), followed by FACS-sorting of mature CD8+ T cells and InEx cells. Subsequently, single-cell TCR (paired ab chains) and RNA sequencing were performed on these cell types. An HLA-B27+ reactive arthritis (ReA) sample were used as a comparison. These were all compared to HLA-B27+ healthy controls (n=4). Additionally, we used the GLIPH2 computational tool to assess TCR sequence convergence based on shared motifs to infer antigen-specific TCR groups.

Results: 12 distinct clusters of mature CD8+ T cells from all samples were observed, exhibiting gene expression consistent with TRM-like, cytotoxic and interferon-stimulated gene (ISG) potential. axSpA InEx cells showed a preferential expression of clusters associated with ISG expression and proliferation vs ReA mature CD8+ T cells, as judged by high IFIT1, IFIT3, MX1, and MKI67 transcripts. Increased cytotoxic potential, based on elevated GZMA and GZMB, with some suppressive capacity, based on elevated IL2RA, FOXP3, and CTLA4, was also observed in axSpA InEx cells. The axSpA InEx TCR repertoire was less diverse, clonally expanded, and demonstrated specific TRBV9_TRAV27, TRBV9_TRAV8-2, and TRBV7-8_TRAV3 gene usage vs mature CD8+ T cells from either axSpA PB or SF. However, axSpA InEx cells demonstrated similarities with ReA mature CD8+ T cells, based on TCR gene usage (TRBV20-1_TRAV1-2, TRBV6-1_TRAV1-2, and TRBV6-2_TRAV1-2) and CDR3b motifs.

Conclusion: These observations suggest that InEx cells may incite inflammation similarly to ReA due to potential recognition of similar peptides, ultimately driving the selection of specific T cell clones. However, InEx cells represent a hybrid tissue resident memory-like population, likely primed by chronic type I interferon exposure. This may underlie the functional capacity of their clonal expansion and their ability to perpetuate inflammation which is distinct from ReA. This has important implications for therapeutic designs attempting to target integrin blockade in axSpA.

Left panel displaying transcriptome profile of InEx vs mature CD8+ ReA SFMCs on UMAP plots. Genetic profile of InEx cells cluster differently to ReA SF. Right panel displaying notable gene expression across all groups, particularly in InEx and ReA SFMC. UMAP = Uniform Manifold Approximation and Projection, SF = synovial fluid, HC = healthy controls.

Left panel displaying the V gene frequency (% total cells in the group) of paired TRAV_TRBV genes expanded in InEx cells vs axSpA PBMC. Right panel displaying the CDR3b sequence logo plots between InEx and ReA SF mature CD8 T cells. Yellow stars highlight the amino acids that differ between the two groups.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/multi-omic-profiling-of-cd8-t-cells-in-axial-spondyloarthritis-axspa-and-reactive-arthritis-rea-implicates-common-pathways/