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Mildly oxidized low-density lipoprotein activates platelets through lysophosphatidic acid (LPA). Hence, the platelet-activating properties attributed to native low-density lipoprotein (nLDL) might be caused by LPA contamination. We show that nLDL enhances thrombin receptor-activating peptide (TRAP)-induced fibrinogen binding to KIIbL L3. The LPA receptor blocker N-palmitoyl-L-serine-phosphoric acid did not affect nLDL-enhanced fibrinogen binding induced by TRAP, but reduced TRAP-induced binding. cAMP and inhibitors of protein kinase C and Ca 2+ rises completely blocked ligand binding by TRAP and nLDL/TRAP. Inhibitors of p38 MAPK and ADP secretion interfered only partially. Blockade of Rho-kinase increased ligand binding 2^3-fold. We conclude that nLDL enhances TRAP-induced fibrinogen binding independent of LPA. fl 2001 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.

Objective— Scavenger receptor BI (SR-BI) is a cell surface receptor that promotes the selective uptake of cholesteryl esters from high-density lipoprotein (HDL) by the liver. In mice, SR-BI deficiency results in increased plasma HDL cholesterol levels and enhanced susceptibility to atherosclerosis. The aim of this study was to investigate the role of SR-BI deficiency on platelet function. Methods and Results— SR-BI-deficient mice were thrombocytopenic, and their platelets were abnormally large, probably because of an increased cholesterol content. The FeCl 3 acute injury model to study arterial thrombosis susceptibility showed that SR-BI wild-type mice developed total arterial occlusion after 24±2 minutes. In SR-BI-deficient mice, however, the time to occlusion was reduced to 13±1 minutes ( P =0.02). Correspondingly, in SR-BI-deficient mice, platelets circulated in an activated state and showed increased adherence to immobilized fibrinogen. In contrast, platelet-specific disruption of SR-BI by bone marrow transplantation in wild-type mice did not alter plasma cholesterol levels or affect platelet count, size, cholesterol content, or reactivity, suggesting that changes in plasma cholesterol levels were responsible for the altered responsiveness of platelets in SR-BI-deficient mice. Conclusion— The function of SR-BI in HDL cholesterol homeostasis and prevention of atherosclerosis is indirectly also essential for maintaining normal platelet function and prevention of thrombosis.

The interaction of platelets with low-density lipoprotein (LDL) plays an important role in the pathogenesis of atherosclerosis and thrombosis. Previously, we have shown that native LDL (nLDL) is a mild activator of platelets increasing their sensitivity to aggregation-inducing agents. Binding of nLDL to platelets was saturable, reversible and initiated signal transduction to p38MAPK suggesting the involvement of a receptor. A peptide mimic of the B-site in apoB100 resembled nLDL in its platelet-activating properties, suggesting that the receptor is a member of the LDL-receptor family. Platelets from familial hypercholesterolemia patients, who lack or have a defective apoB/E receptor, responded normally to nLDL and an antibody against this receptor left nLDL-induced activation of normal platelets undisturbed, excluding the involvement of the classical LDL (apoB/E)-receptor. In this study, we provide evidence that nLDL initiates platelet signaling to p38MAPK via a splice variant of the LDL-receptor family member Apolipoprotein E Receptor 2 (ApoER2′). This conclusion is based on (i) blockade of nLDL-induced p38MAPK activation by receptor-associated protein (RAP), an inhibitor of ligand binding to members of the LDL-receptor family, (ii) confocal microscopy showing a high degree of co-localization of nLDL and ApoER2′ at the platelet surface, (iii) binding of both nLDL and the B-site peptide to soluble ApoER2′, (iv) activation of p38MAPK by an anti-ApoER2 antibody with similar kinetics as nLDL, and (v) tyrosine phosphorylation of ApoER2′ upon binding of nLDL. The nLDL-induced phosphorylation of ApoER2′ could be abolished by PP1, an inhibitor of Src-like tyrosine kinases. In the absence of PP1, ApoER2′ phosphorylation was accompanied by co-association with the Src-family member Fgr. We conclude that binding of nLDL to platelets involves ApoER2′. Upon nLDL binding, the receptor is phosphorylated which induces the recruitment of Fgr, a kinase known to activate p38MAPK. The ApoER2′-Fgr complex subsequently activates p38MAPK, an upstream element in the formation of thromboxane A2 that primes the platelets to further stimulation by aggregation-inducing agents.