Heparin modulates integrin function in human platelets

AbstractPurpose: Heparin binds to human platelets and can cause activation and aggregation, although the mechanisms are unknown. To determine how heparin alters platelet function, we identified platelet-binding sites for heparin and measured heparin's influence on the function of platelet integrin αIIbβ3 (glycoprotein IIb/IIIa). Methods: Photoaffinity cross-linking and affinity chromatography experiments were performed to identify platelet membrane proteins that bind heparin. Heparin's effect on fibrinogen binding to platelets was measured with a radioligand-binding assay. The translocation to the cytoskeleton of Rap2, a guanosine triphosphate–binding protein, was measured from platelets aggregating in response to heparin and other agonists. Results: Cross-linking and affinity chromatographic experiments positively identified the integrin αIIbβ3 as a heparin-binding site. Heparin aggregation was calcium dependent. Low concentrations of unfractionated porcine mucosal heparin (2-5 U/mL) significantly increased fibrinogen I 125 binding to activated platelets, whereas higher doses did not. Heparin-mediated platelet aggregation was completely blocked by GRGDS peptide (5 mmol/L), a competitive inhibitor of fibrinogen binding, and was blocked by EDTA (2 mmol/L), which dissociates the functional integrin complex. Aggregation was associated with Rap2 translocation to the cytoskeleton, a sign of outside-in signaling. Conclusions: Heparin binds to the αIIbβ3 integrin in vitro and ex vivo, and heparin increases fibrinogen binding to the integrin. Heparin-mediated aggregation requires an intact integrin and ligand and leads to Rap2 translocation to the cytoskeleton—an outside-in signal of ligand engagement. Heparin may directly modulate platelet integrin function, most likely through direct binding and modulation of integrin function. (J Vasc Surg 2001;33:587-94.