THU0055 An anatomically distinct pathogenic fibroblast subset drives inflammation in arthritis
Authors: A.P. Croft, J. campos, J. Marshall, J. Turner, F. Barone, A. Filer, C. Buckley
Journal: Annals of the Rheumatic Diseases (2018)

<h3>Background</h3> Fibroblasts are key effector cells in the persistence of synovial inflammation and joint damage. It is not yet known whether specific subsets of synovial fibroblasts exist, and if so, if they are responsible for the distinct fibroblast mediated features observed in inflammatory arthritis, such as invasion of cartilage, bone damage, and persistence of inflammation. <h3>Objectives</h3> Here we identify and describe the biology of a functionally distinct pathogenic fibroblast cell type marked by the co-expression of Thy1.2 and Podoplanin (Pdpn), that is responsible for persistence of synovial inflammation. <h3>Methods</h3> We used the serum transfer arthritis (STA) model to induce joint inflammation. To identify putative subsets of fibroblasts we used flow cytometry using established markers of stromal cells, on enzymatically digested synovial tissue. These cells were localised in tissue sections using immunohistochemistry or immunofluorescence staining. Proliferation of subsets <i>in vivo</i> during the time course of STA was determined by BrdU incorporation studies. Transcriptomic analysis was performed using ultra-low input RNA sequencing on flow sorted populations of cells and single cell analysis performed on CD45 negative gated cells. <h3>Results</h3> Using FAP as a biomarker of activated synovial fibroblasts we have been able to identify and define distinct subsets of synovial fibroblasts based on their co-expression of Thy1.2 and Pdpn. These subsets reside in distinct compartments of the synovial microanatomy including the lining layer (LL), sub-lining layer (SL) and a subset of pericytes. We found that FAP +cells within the SL are highly proliferative and their expansion in cell number positively correlates with inflammation. Global deletion of FAP +mesenchymal cells in the synovium using a FAP-DTR mouse attenuated synovial inflammation; protected against erosive bone damage and lead to reduced leucocyte accumulation as a result of reduced chemokine and cytokine production by synovial fibroblasts within the membrane. Collectively these data suggest a pathogenic pro-inflammatory role for these cells. Transcriptomic analysis by both ultra-low input bulk RNA sequencing and single cell analysis revealed these subsets to be transcriptionally distinct with the greatest transcriptional differences observed between LL and SL cells, demonstrating a site specific transcriptional program for cells within these compartments. Gene signature analysis of SL FAP +cells was consistent with an immune effector cell phenotype, in contrast to lining layer cells that express genes associated with matrix remodelling. Finally, to test this hypothesis we injected FAP +LL cells and FAP +SL cells into inflamed ankle joints of mice. The injection of FAP +SL cells lead to more severe and prolonged joint inflammation, whereas injection of LL FAP +cells had no effect. Finally, we identified these cell subsets within human synovial tissue and have demonstrated the expansion of SL cells with inflammation. <h3>Conclusions</h3> Synovial inflammation is associated with the expansion, activation and differentiation of fibroblasts into distinct functional subsets of cells that regulate those specific aspects of inflammatory joint pathology. Direct targeting of specific pathogenic subsets of synovial fibroblasts may provide a novel, non-immunosuppressive approach to the treatment of inflammatory arthritis. <h3>Disclosure of Interest</h3> None declared