Background/Purpose: Effective treatment of Rheumatoid arthritis (RA) patients is achievable within a short window of opportunity following diagnosis. Identification of pathogenic immune mechanisms at a pre-RA stage would greatly benefit our understanding of the early events that govern disease progression and help identify early points of therapeutic intervention.

Characterization of pro-inflammatory T cell polyfunctionality in the periphery and synovial tissue of ’at-risk; subjects (Arthralgia) RA patients (RA) and healthy controls (HC).

Identification of pathogenic, synovial T cell subsets.

Characterization of metabolic status of pathogenic T cell populations.

Methods: Synovial biopsies from RA, AR and HC were obtained by arthroscopic surgery followed by RNAseq analysis (Guo et al., PLoS One, 2018). Single cell synovial tissue cell suspensions from RA, AR and HC and paired PBMC were stimulated in vitro and polyfunctional synovial T cell subsets examined by flow cytometric analysis, SPICE visualization and Flowsome unsupervised clustering. Flow-Imaging was utilised to confirm specific T cell cluster identification. Fluorescent Lifetime Imaging Microscopy (FLIM) was used to visualise metabolic status of specific T cell populations.

Results: T cell associated pro-inflammatory cytokines were increased in whole synovial tissue biopsy RNAseq analysis in RA patients and AR subjects compared to HCs. Flow cytometric analysis and SPICE revealed marked T cell polyfunctionality with similar pro-inflammatory cytokine profiles in RA and AR compared to HC, providing evidence of a dysregulated synovial T cell response that pre-dates clinical onset of disease. Cluster analysis led to the identification of highly polyfunctional T cell clusters and an accumulation of CD4⁺CD8⁺ double positive T cells in the synovial tissue of AR and RA but not in HC. Hybrid flow cytometry and imaging technique confirmed the cell membrane co-expression of CD4 and CD8 by a synovial T cell population. DP T cells do not share characteristics of NK T cells and their synovial accumulation correlates with DAS28(CRP). Initial studies utilising non-invasive FLIM technique and flow cytometric analysis revealed that DP T cells have an intermediate metabolic profile between CD4 and CD8 T cells and an activated mTOR pathway.

Conclusion: Highly polyfunctional pro-inflammatory T cell responses pre-date disease onset as demonstrated by the accumulation of polyfunctional T cells in the synovial tissue of pre-RA arthralgia subjects. These data highlight a key early pathogenic role for T cell plasticity and specific synovial T cell clusters including, DP T cells in RA.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/highly-polyfunctional-metabolically-altered-pathogenic-t-cells-accumulate-in-the-synovial-tissue-of-ra-patients-and-arthralgia-subject-but-not-healthy-control-synovial-tissue/