Systemic sclerosis sera affect fibrillin-1 deposition by dermal blood microvascular endothelial cells: therapeutic implications of cyclophosphamide

Abstract INTRODUCTION: Systemic sclerosis (SSc) is a connective tissue disorder characterized by endothelial cell injury, autoimmunity and fibrosis. Fibrillin-1 alterations were reported in SSc: i) fibrillin-1 microfibrils are disorganized in SSc dermis, ii) fibrillin-1 microfibrils produced by SSc fibroblasts are unstable, iii) mutations in FBN1 gene and anti-fibrillin-1 autoantibodies were reported in SSc. Fibrillin-1 microfibrils, which are abundantly produced by blood and lymphatic microvascular endothelial cells (B-MVECs, Ly-MVECs), sequester in the extracellular matrix the latent form of the potent profibrotic cytokine transforming growth factor- (TGF-beta). In the present study we evaluated the effects of SSc sera on the deposition of fibrillin-1 and microfibril-associated glycoprotein-1 (MAGP-1) and the expression of focal adhesion molecules by dermal B-MVECs and Ly-MVECs. METHODS: Dermal B-MVECs and Ly-MVECs were challenged with sera from SSc patients, naive or under cyclophosphamide (CYC) treatment, and healthy controls. Fibrillin-1/MAGP-1 synthesis and deposition and the expression of alphavbeta3 integrin/phosphorylated-FAK and vinculin/phalloidin were evaluated by immunofluorescence and quantified by morphometric analysis. RESULTS: Fibrillin-1 and MAGP-1 colocalized in all experimental conditions, forming a honeycomb pattern in B-MVECs and a dense mesh of short segments in Ly-MVECs. In B-MVECs, fibrillin-1/MAGP-1 production and alphavbeta3 integrin expression significantly decreased upon challenge with sera from naive SSc patients compared with healthy controls. Upon challenging of B-MVECs with sera from CYC-treated SSc patients, fibrillin-1/MAGP-1 and alphavbeta3 integrin levels were comparable to those of cells treated with healthy sera. Ly-MVECs challenged with SSc sera did not differ from healthy controls in the expression of any of the molecules assayed. CONCLUSIONS: Due to the critical role of fibrillin-1 in sequestering the latent form of TGF-beta in the extracellular matrix, its decreased deposition by B-MVECs challenged with SSc sera might contribute to dermal fibrosis. In SSc, CYC treatment might limit fibrosis through the maintenance of physiological fibrillin-1 synthesis and deposition by B-MVECs.