HEPARIN BINDING TO HUMAN MONOCYTES: MODULATION BY HISTIDINE-RICH GLYCOPROTEIN

Abstract

Heparin and its related glycosaminoglycans interact with a variety of cell types and, irrespective of their anticoagulant activities, have a complex and biologically important influence in blood vessel wall biology. In this study, the binding of heparin to the human monocytoid cell line U937 was characterized. 35s-Heparin bound to U937 cells in a specific, concentration-dependent, and saturable manner, while binding to human erythrocytes was minimal. The binding was rapid, reaching a steady state at 30 min at 4°C and was reversible. Excess unlabeled heparin, but not chondroitin sulfate or dermatan sulfate, inhibited heparin binding, demonstrating the specificity for heparin. Scatchard plot analysis revealed a single class of heparin binding sites, with an apparent Kd of 0.32 uM and approx. 2.0 × 106 sites/cell. When binding was performed at 37°C, significant amount of cell-bound heparin was not removed by cell surface trypsinization, indicating endocytosis of heparin by U937 cells. Taken together the binding characteristics suggested the presence of specific heparin receptors on U937 cell surface. Histidine-rich glycoprotein (HRGP), a potent heparin-binding protein found in human plasma and platelets, decreased the affinity of heparin for U937 cells without a significant change in maximal binding. This negative modulation of heparin binding was specific for HRGP since antithrombin III was much less effective. While EDTA abolished the HRGP-heparin interaction, it had no effect on the HRGP modulation of heparin cell binding, suggesting that HRGP interacted directly with the U937 cell and altered its affinity for heparin. Specific binding of labeled HRGP to U937 cells was demonstrated. Cell-surface bound heparin markedly accelerated the inactivation of thrombin by antithrombin III. Its anticoagulant potency was equivalent to heparin in the fluid phase. Human monocytes and macrophages provide an optimal cell surface for the assembly of prothrombinase and the generation of thrombin. The specific binding of anticoagulantly active heparin to monocytes and macrophages may modulate the procoagulant properties of these cells at sites of inflammation and thrombosis. Furthermore, since heparin fractions with high or low affinity for antithrombin III bound equally well to U937 cells, heparin may have a direct effect on monocyte cell biology. The current study also raises the possibility that the significant modulation of heparin cell binding by HRGP may represent a major function for this protein in vivo.