SAT0193 Macrophages Responding To Mechanical Stress in Scleroderma

Background Inflammation, vasculopathy and tissue fibrosis are key features of scleroderma (SSc). While healthy forearm skin which has a Young9s modulus of 4–12kPa, SSc fibrotic skin measures 50–80kPa with its increased mechanical stiffness 1 . We have shown that mechano-sensing properties of fibroblasts in SSc are mediated by myocardin-related transcription factor A (MRTF-A) 3 . Monocytes/macrophages are likely to be involved in SSc pathogenesis but the effect of mechanical stress on these cells remain to be elucidated 3,4 . Objectives To investigate if a mechanically-stressed microenvironment like that in SSc tissue, promotes macrophages towards a pathogenic phenotype, and whether MRTF-A is involved in this process. Methods Control and scleroderma skin sections were immunostained with anti-CD68 and anti-MRTF-A antibodies (n=3). Human PBMC-derived macrophages were cultured in RPMI/M-CSF on 4kPa and 50kPa collagen-fibronectin-coated plates to mimic “soft”/healthy and “stiff”/fibrotic skin, and activated with LPS (10ng/ml) or IL-10 (10ng/ml) for macrophages designated M(LPS) and M(IL-10) (n=4). MRTF-A expression was assessed by qPCR and conditioned media profiled by Luminex array for inflammatory cytokines. Mouse bone marrow-derived macrophages (BMDMs) of wildtype and MRTF-A-null mice were maintained in RPMI/M-CSF on soft and stiff substrates. The data were analysed by two-way ANOVA and Tukey test (p Results We observed a greater number of CD68 + macrophages in diffuse SSc skin compared to control skin, mainly around perivascular regions. These macrophages also expressed MRTF-A. Human macrophages expressed MRTF-A mRNA and showed differential cytokine expression when cultured on soft and stiff substrates. M(LPS) on soft matrix expressed IFN-γ, which was undetectable with M(LPS) on stiff substrate (mean difference 0.2075±0.1576pg/ml, p Conclusions MRTF-A is a mechanical stress-responsive factor which co-activates transcription of cytoskeletal and extracellular matrix-modifying genes. MRTF-A may couple mechanical stress to macrophage activation in SSc, where stiff matrix promotes macrophage secretion of cytokines and growth factors that exacerbate fibrosis. References Sacksen et al., Arthritis Rheum 2013. Shiwen et al., PLoS One 2015. Stifano et al., Curr Rheumatol Rep 2015. Chia et al., Curr Opin Rheumatol 2015. Disclosure of Interest None declared