Abstract 555: Bone Morphogenetic Protein Modulator BMPER Induces Angiogenesis via Inhibition of Thrombospondin-1 and Activation of the Fibroblast Growth Factor Signaling Pathway

Abstract

Introduction: Bone morphogenetic proteins (BMP) play a key role in vascular development. Previously, we have identified BMP endothelial cell precursor-derived regulator (BMPER), an extracellular BMP modulator, to increase the angiogenic activity of endothelial cells in a concentration-dependent manner. In this project we now investigate how the BMPER effect is mediated by key molecules of angiogenesis. Methods and Results: To assess the effect of BMPER on angiogenesis-related molecules we performed an angiogenesis antibody array with BMPER-stimulated human umbilical venous endothelial cells (HUVECs) and vice versa with BMPER-silenced HUVECs compared to control conditions, respectively. We detected increased protein expression of the anti-angiogenic thrombospondin-1 (TSP-1) 48 hours after siBMPER transfection and, consistently, decreased TSP-1 expression after stimulation with BMPER (60 ng/ml; 39% ± 7.3 N=4). Furthermore, the pro-angiogenic protein bFGF was increased after BMPER stimulation, which was confirmed by realtime-PCR and western blot analysis (288.8% ± 74.8 N=3). Additionally, we detected increased FGF receptor-1 protein expression (137.7% ± 0.4 N=3) as well as FGF signaling pathway activation. Next, we investigated the interaction of BMPER and the FGF signaling pathway in endothelial cell function. BMPER stimulation increased HUVEC angiogenic activity in matrigel, migration and spheroid assays and concomitant inhibition of FGF signaling by an anti-bFGF antibody effectively inhibited the pro-angiogenic BMPER effect. Silencing of BMPER decreased the expression of FGFR1 and, accordingly, stimulation of BMPER-silenced cells with bFGF showed decreased angiogenic endothelial cell activity (65%) compared to control. The angiogenic activity of bFGF was also reduced in C57BL/6_Bmper +/- mice as assessed in the matrigel plug assay. Ex vivo aortic ring assays of C57BL/6_Bmper +/- mice confirmed a specific effect for bFGF but not for VEGF. Conclusion: In summary, BMPER inhibits the expression of the anti-angiogenic TSP-1 and increased the expression as well as activation of the pro-angiogenic FGF signaling pathway, which overall lead to the promotion of angiogenesis.