Decorin Is a Novel VEGFR-2-Binding Antagonist for the Human Extravillous Trophoblast

Extravillous trophoblasts (EVT) of the human placenta invade the uterine decidua and its arteries to ensure successful placentation.. We previously identified two decidua-derived molecules, TGF-beta and a TGF-beta-binding proteoglycan decorin (DCN), as negative regulators of EVT proliferation, migration, and invasiveness and reported that DCN acts via multiple tyrosine kinase receptors [epidermal growth factor-receptor (EGF-R), IGF receptor-1 (IGFR1), and vascular endothelial growth factor 2 receptor (VEGFR-2)]. Because binding of DCN to VEGFR-2 has never been reported earlier, present study explored this binding, the approximate location of VEGFR-2-binding site in DCN, and its functional role in our human first trimester EVT cell line HTR-8/SVneo. Based on far-Western blotting and coimmunoprecipitation studies, we report that DCN binds both native (EVT expressed) and recombinant VEGFR-2 and that this binding is abrogated with a VEGFR-2 blocking antibody, indicating an overlap between the ligand-binding and the DCN-binding domains of VEGFR-2. We determined that I-125-labeled VEGF-E (a VEGFR-2 specific ligand) binds EVT with a dissociation constant (K-d) of 566 pM, and DCN displaced this binding with an inhibition constant (K-i) of 3.93-5.78 nM, indicating a 7- to 10-fold lower affinity of DCN for VEGFR-2. DCN peptide fragments derived from the leucine rich repeat 5 domain that blocked DCN-VEGFR-2 interactions or VEGF-E binding in EVT cells also blocked VEGF-A- and VEGF-E-induced EVT cell proliferation and migration, indicative of functional VEGFR-2-binding sites of DCN. Finally, DCN inhibited VEGF-E-induced EVT migration by interfering with ERK1/2 activation. Our findings reveal a novel role of DCN as an antagonistic ligand for VEGFR-2, having implications for patho-physiology of preeclampsia, a trophoblast hypoinvasive disorder in pregnancy, and explain its antiangiogenic function. (Molecular Endocrinology 25: 1431-1443, 2011)