Inside the Joint of Inflammatory Arthritis Patients: Handling and Processing of Synovial Tissue Biopsies for High Throughput Analysis

Abstract

Inflammatory arthritis is a chronic systemic autoimmune disease of unknown etiology, which affects the joints. If untreated, these diseases can have a detrimental effect on the patient's quality of life, leading to disabilities, and therefore, exhibit a significant socioeconomic impact and burden. While studies of immune cell populations in arthritis patient's 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. Understanding the contribution of immune and stromal cell interactions within the inflamed joint has been met with several technical challenges. Additionally, the limited availability of synovial tissue biopsies is a key incentive for the utilization of high-throughput techniques in order to maximize information gain. This review aims to provide an overview of key methods and novel techniques that are used in the handling, processing and analysis of synovial tissue biopsies and the potential synergy between these techniques. Herein, we describe the utilization of high dimensionality flow cytometric analysis, single cell RNA sequencing, ex vivo functional assays and non-intrusive metabolic characterization of synovial cells on a single cell level based on fluorescent lifetime imaging microscopy. Additionally, we recommend important points of consideration regarding the effect of different storage and handling techniques on downstream analysis of synovial tissue samples. The introduction of new powerful techniques in the study of synovial tissue inflammation, brings new challenges but importantly, significant opportunities. Implementation of novel approaches will accelerate our path toward understanding of the mechanisms involved in the pathogenesis of inflammatory arthritis and lead to the identification of new avenues of therapeutic intervention.

Computational approaches in rheumatic diseases – Deciphering complex spatio-temporal cell interactions

“…Very early implementation of flow cytometric analysis of cultured synovial fibroblasts isolated from patient with RA synovial biopsies had limited capacity but showed differences in autofluorescence that reflect morphological changes and mitochondrial activity of RA derived compared to healthy control derived synovial fibroblasts [36] . Despite challenges in sample preparation, more recent implementation of mass cytometry has eluted to multiple and functionally distinct synovial fibroblast subsets complementing transcriptomic differences identified by scRNAseq [37] , [38] , [39] , [40] . Flow cytometric analysis has also been extensively utilised for the study of patient with IA synovial T cell responses resulting in the identification of pathogenic CD161 + ex-Th17 cells that show increased polyfunctionality due to the secretion of multiple pro-inflammatory cytokines simultaneously and resistance to autologous Treg cell suppression [41] , [42] .…”