FRI0342 Degranulation of Mast Cells is Enhanced by Direct Contact with Rheumatoid Arthritis Synovial Fibroblasts but not Dermal Fibroblasts
Authors: E.Y. Kim, J.M. Lee, Y.W. Song, K. Shin
Journal: Annals of the Rheumatic Diseases (2014)

<h3>Background</h3> A number of studies have shown that synovial mast cells are activated, and elicit a pro-inflammatory role in rheumatoid arthritis (RA). Synovial mast cells are located around the microvasculature, but also adjacent to the synovial lining and subsynovium where synovial fibroblasts (SF) are residing. Despite that mediators produced by mast cells have been demonstrated to activate SF, the effector function of SF on mast cell activation is not well known. <h3>Objectives</h3> To study the whether mast cells could be activated upon direct or indirect contact with RA SF in vitro <h3>Methods</h3> In vitro SF cultures were obtained from knee synovium of RA patients. Dermal fibroblasts from healthy individuals was used for comparison. Co-culture of SF (2.5x10<sup>4</sup> cells) and human mast cell (HMC)-1 line (ratio 1:1, 1:2, 1:5) was done with IMEM phenol red free media including 10% fetal bovine serum in a 24 well plate. We used the hexosaminidase (HXSA) assay as the read-out for mast cell degranulation (1, 6, 12, 24 hours) after the co-culture. To exempt the effect of HXSA release by RA SF and mast cell alone, control cultures were also performed in parallel. A transwell system - mast cells were cultured in the upper well insert - was used to compare results of direct vs. indirect contact between RA SF and HMC-1. Each experiment was repeated at least three times independently with different RA SF batches. <h3>Results</h3> The RA SF-HMC co-culture was well maintained without requiring special additives or conditions; majority of mast cells were in direct contact of SF. HXSA release was enhanced in direct co-culture conditions at 24 hours (Figure A). However, minimal if any additional release was detected in indirect co-cultures, implying that direct RA SF-mast cell interaction can induce mast cell activation. Increased HXSA release in direct co-cultures was appreciated after 12 hours of co-culture, which shows that the results are not driven solely by an immediate release by RA SF ligand-mast cell receptor interaction. Of note, HMC-1 co-culture with dermal fibroblasts at 24 hours only resulted in mild increase of HXSA release compared with RA SF-HMC co-cultures (Figure B). <h3>Conclusions</h3> Our results demonstrate that there may be a cross-talk between SF and mast cells in proximity, resulting in boosted degranulation of mast cells within the rheumatoid synovium. <h3>References</h3> Skin K, Nigrovic PA, Crish et al. Mast Cells Contribute to Autoimmune Inflammatory Arthritis via Their Tryptase/Heparin Complexes. J Immunol 2009;182:647-56. Keida S, Shin K, Nigrovic PA et al. Synovial Fibroblasts Promote the Expression and Granule Accumulation of Tryptase via Interleukin-33 and Its Receptor ST-2 (IL1RL1). J Biol Chem 2010;28:21478-86. <h3>Acknowledgements</h3> We would like to thank Dr. Butterfield for providing the HMC-1 cell line and Dr. Jinho Chung for the dermal fibroblasts. <h3>Disclosure of Interest</h3> None declared <h3>DOI</h3> 10.1136/annrheumdis-2014-eular.4850