Human umbilical vein endothelial cells express high affinity receptors for factor Xa

Abstract

The binding of [125I]-factor Xa to human umbilical vein endothelial cell (HUVEC) monolayers was studied. At 7 degrees C, [125I]-factor Xa bound to a single class of binding sites with a dissociation constant value of 6.6 +/- 0.8 nM and a binding site density of 57,460 +/- 5,200 sites/cell (n = 3). Association and dissociation kinetics were of a pseudo-first order and gave association and dissociation rate constant values of 0.15 x 10(6) M-1 s-1 and 4.0 x 10(-4) s-1, respectively. [125I]-factor Xa binding was inhibited by factor Xa but was not affected by factor X, thrombin or monoclonal antibodies against factor V, antithrombin-III or tissue factor pathway inhibitor (TFPI) but was inhibited by an antibody specific for the effector cell protease receptor-1 (EPR-1), a well-known receptor of factor Xa on various cell types. [125I]-factor Xa binding to HUVEC was not affected by various inhibitors of factor Xa such as DX 9065, pentasaccharide-antithrombin-III or TFPI. Factor Xa increased intracellular free calcium levels and phosphoinositide turnover in endothelial cells and, when added to HUVEC in culture, factor Xa was a potent mitogen, stimulating an increase in cell number at a 0.3 to 100 nM concentration. HUVEC-bound factor Xa promoted prothrombin activation in the presence of factor Va only. This effect was inhibited by both indirect and direct inhibitors of factor Xa. These findings indicate that HUVEC express functional high affinity receptors for factor Xa, related to EPR-1, which may be of importance in the regulation of coagulation and homeostasis of the vascular wall.

Regulatory Mechanism of Matrix Metalloprotease-2 Enzymatic Activity by Factor Xa and Thrombin

“…Factor Xa acts as a potent mitogen for endothelial cells (23), fibroblasts (24), and vascular SMCs (25,26). Both proteases can also elicit endothelial cell and SMC migration through pro-MMP-2 activation and subsequent extracellular matrix degradation (13,27,28).…”

Coagulation Factors VIIa and Xa Induce Cell Signaling Leading to Up-regulation of the egr-1 Gene

“…In endothelial cells addition of FXa can activate NO synthase (13,14) and induce synthesis of cytokines and expression of adhesion molecules (14,15). Antibodies to PDGF (11,16,17) or effector cell protease receptor-1 (EPR-1) (17,18) both attenuate mitogenic responses to FXa in endothelial cells and in vascular smooth muscle cells. Factor V (FV), the cellular cofactor for FXa in formation of the prothrombinase complex (2), has to our knowledge not been shown to participate in any of these events.…”

Factor Xa Activates Endothelial Cells by a Receptor Cascade Between EPR-1 and PAR-2

“…This protein, called effector protease receptor-1 (EPR-1), behaves as a cofactor for factor Xa to catalyze prothrombin activation in the absence of added factor Va. 2 Recently, we and others demonstrated the existence of such a population of high-affinity, functional, factor Xa-binding sites in human vascular endothelial cells (HUVECs) and showed that exposure of HUVECs to factor Xa induced phosphoinositide turnover and an increase in intracellular free Ca 2ϩ . 3,4 Most important, through binding to this receptor, factor Xa was also a potent mitogen for endothelial cells. 3 Moreover, a recent article by Papapetropoulos et al 5 showed increased interleukin-6 release by HUVECs after treatment with factor Xa.…”

“…3,4 Most important, through binding to this receptor, factor Xa was also a potent mitogen for endothelial cells. 3 Moreover, a recent article by Papapetropoulos et al 5 showed increased interleukin-6 release by HUVECs after treatment with factor Xa. These activities of factor Xa are affected by selective factor Xa inhibitors, which suggests that occupancy of EPR-1 alone by factor Xa is not sufficient for cell activation and that protease activity is required for factor Xa to induce signal transduction and subsequent activation of vascular endothelial cells.…”