Background/Purpose: Inflammatory arthritis (IA) is a chronic systemic autoimmune disease of unknown aetiology, which affects the joints. While studies of immune cell populations in peripheral blood have been informative regarding potential immune cell dysfunction and possible patient stratification, there are considerable limitations in identifying the early events that lead to synovial inflammation. The joint, as the site of inflammation and the local microenvironment, exhibit unique characteristics that contribute to disease pathogenesis. This work aims to provide an overview of high throughput analysis and key methodology for the handling, processing and analysis of ST biopsies and the potential synergy between these techniques.

Methods: Synovial biopsies were obtained from inflammatory arthritis (IA) patients at arthroscopy (St. Vincent’s University Hospital). Paired PBMC were obtained on the same day. Methodology included: flow cytometric analysis, RNA sequencing, ex vivo functional assays (T cell activation and endocytosis). Ethics for this study was approved by the St. Vincent’s University Hospital.

Results: In order to investigate the cellular contribution in the periphery vs the synovial tissue (ST) of IA patients, multi-parameter flow cytometry was utilized. Results suggest that there is an infiltration of specific immune cells into the joint of IA patients, including CD3⁺T cells and myeloid DC, with a specific decrease in B and CD4⁺T cells. When correlating to disease activity (DAS28 and CRP), a specific increase in monocyte-derived cells was observed, including CD14⁺ and CD64⁺ cells. The further stratification between RA and PsA patients, showed specific disease pathotype-infiltration. The study of T cells poly-functionality in IA ST demonstrated a significant decrease in T cells poly-functionality, as shown by the significant decrease in the co-expression of specific cytokines combination (including IL2⁺TNFα⁺GMCSF⁺IFNγ⁺ and GMCSF⁺TNFα⁺ in cells treated with both the OxPhox inhibitor FCCP or the JAK/STAT inhibitor Tofacitinib. In addition, we optimized a new technique to assess endocytosis in multiple populations simultaneously without the need for cell sorting. Briefly, digested cells were incubated in parallel at 4°C (passive endocytosis) and 37°C (active endocytosis) with DQ OVA(Ovalbumin). ST cells were then stained for multiple populations, demonstrating differential endocytosis capacity between RA and PsA ST CD14⁺ and mDC. Finally, utilisation of novel bioinformatics analysis of RNAseq data showed differential gene expression and pathway enrichment involved in IA pathogenesis and allowed for the comparison of cell specific enrichment scores and transcription factor usage based on pathotype and gender.

Conclusion: The introduction of new powerful techniques in the study of ST inflammation, brings new challenges and significant opportunities. These approaches will accelerate our path towards understanding of the mechanisms involved in the pathogenesis of IA and lead to the identification of new avenues of therapeutic intervention.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/high-throughput-analyses-for-the-study-of-synovial-tissue-biopsy-from-inflammatory-arthritis-patients/