Low-Density Lipoprotein Enhances Platelet Secretion Via Integrin-α IIb β 3 –Mediated Signaling

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Abstract
—LDL is known to increase the sensitivity of human platelets for agonists and to induce aggregation and secretion independently at high concentrations, but its mechanism of action is largely obscure. To clarify how LDL increases platelet sensitivity, cells were incubated in lipoprotein-poor plasma and treated with collagen at a concentration that induced ≈20% secretion of 14C-serotonin. Preincubation with LDL (30 minutes at 37°C) enhanced secretion in a dose-dependent manner to 60±14% at a concentration of 2 g LDL protein/L. Similar stimulation by LDL was seen when secretion was induced by the thrombin receptor–activating peptide. This enhancement was strongly reduced (1) in the presence of monoclonal antibody PAC1 against activated αIIbβ3, a polyclonal antibody against αIIb, and in the presence of the fibrinogen peptides GRGDS and HHLGGAKQAGDV; (2) in αIIbβ3-deficient platelets; and (3) after dissociation of αIIbβ3. In contrast, binding of 125I-LDL to normal platelets in the presence of PAC1, anti-αIIb, GRGDS, and HHLGGAKQAGDV, and to αIIbβ3-deficient platelets was normal. LDL increased the surface expression of fibrinogen in lipoprotein-poor plasma and fibrinogen-free medium, suggesting that extracellular and granular fibrinogen bind to αIIbβ3 after platelet-LDL interaction. Platelets deficient in fibrinogen (IIbβ3 preserved responsiveness to LDL, indicating that occupancy of αIIbβ3 was not restricted to fibrinogen. Inhibition of protein kinase C (bisindolylmaleimide) diminished fibrinogen binding and sensitization by LDL; inhibition of tyrosine kinases (herbimycin A) left fibrinogen binding unchanged but diminished sensitization by LDL. We conclude that an increased concentration of LDL, such as observed in homozygous familial hypercholesterolemia, sensitizes platelets to stimulation by collagen and thrombin receptor–activating peptide via ligand-induced outside-in signaling through integrin-αIIbβ3.