title: Impaired platelet activation and cAMP homeostasis in MRP4-deficient mice.
abstract: Molecules that reduce the level of cyclic adenosine 5'-monophosphate (cAMP) in the platelet cytosol, such as adenosine 5'-diphosphate (ADP) secreted from dense granules, trigger platelet activation. Therefore, any change in the distribution and/or availability of cyclic nucleotides or ADP may interfere with platelet reactivity. In this study, we evaluated the role of multidrug resistance protein 4 (MRP4, or ABCC4), a nucleotide transporter, in platelet functions in vivo and in vitro by investigating MRP4-deficient mice. MRP4 deletion resulted in a slight increase in platelet count but had no impact on platelet ultrastructure. In MRP4-deficient mice, the arterial occlusion was delayed and the tail bleeding time was prolonged. In a model of platelet depletion and transfusion mimicking a platelet-specific knockout, mice injected with MRP4(-/-) platelets also showed a significant increase in blood loss compared with mice injected with wild-type platelets. Defective thrombus formation and platelet activation were confirmed in vitro by studying platelet adhesion to collagen in flow conditions, integrin αIIbβ3 activation, washed platelet secretion, and aggregation induced by low concentrations of proteinase-activated receptor 4-activating peptide, U46619, or ADP. We found no role of MRP4 in ADP dense-granule storage, but MRP4 redistributed cAMP from the cytosol to dense granules, as confirmed by increased vasodilator-stimulated phosphoprotein phosphorylation in MRP4-deficient platelets. These data suggest that MRP4 promotes platelet aggregation by modulating the cAMP-protein kinase A signaling pathway, suggesting that MRP4 might serve as a target for novel antiplatelet agents.
authors: INSERM Unité Mixte de Recherche S1140, Faculté de Pharmacie, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, Paris, France;
authors: Benoit
authors: B
authors: Decouture
authors: INSERM Unité Mixte de Recherche S1140, Faculté de Pharmacie, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, Paris, France;
authors: Elise
authors: E
authors: Dreano
authors: INSERM Unité Mixte de Recherche S1140, Faculté de Pharmacie, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, Paris, France; Assistance Publique-Hôpitaux de Paris, Hôpital Européen Georges Pompidou, Service d'Hématologie Biologique, Paris, France;
authors: Tiphaine
authors: T
authors: Belleville-Rolland
authors: INSERM Unité Mixte de Recherche S1140, Faculté de Pharmacie, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, Paris, France;
authors: Orjeta
authors: O
authors: Kuci
authors: INSERM Unité Mixte de Recherche S1140, Faculté de Pharmacie, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, Paris, France;
authors: Blandine
authors: B
authors: Dizier
authors: INSERM Unité Mixte de Recherche S1176, Université Paris-Sud, Le Kremlin-Bicêtre, France;
authors: Amine
authors: A
authors: Bazaa
authors: Université Paris Descartes, Sorbonne Paris Cité, Paris, France; Equipe d'Accueil 4475, Faculté de Pharmacie, Paris, France;
authors: Bérard
authors: B
authors: Coqueran
authors: Sorbonne Universités, Université Pierre et Marie Curie Univ Paris 06, Faculté de Médecine, Unité Mixte de Recherche S1166, Institute of Cardiometabolism and Nutrition, Paris, France; and Assistance Publique-Hôpitaux de Paris, Hôpital Pitié-Salpêtrière, Service de Pharmacologie et Centre d'Investigation Clinique 1421, Paris, France.
authors: Anne-Marie
authors: AM
authors: Lompre
authors: INSERM Unité Mixte de Recherche S1176, Université Paris-Sud, Le Kremlin-Bicêtre, France;
authors: Cécile V
authors: CV
authors: Denis
authors: Sorbonne Universités, Université Pierre et Marie Curie Univ Paris 06, Faculté de Médecine, Unité Mixte de Recherche S1166, Institute of Cardiometabolism and Nutrition, Paris, France; and Assistance Publique-Hôpitaux de Paris, Hôpital Pitié-Salpêtrière, Service de Pharmacologie et Centre d'Investigation Clinique 1421, Paris, France.
authors: Jean-Sébastien
authors: JS
authors: Hulot
authors: INSERM Unité Mixte de Recherche S1140, Faculté de Pharmacie, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, Paris, France;
authors: Christilla
authors: C
authors: Bachelot-Loza
authors: INSERM Unité Mixte de Recherche S1140, Faculté de Pharmacie, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, Paris, France; Assistance Publique-Hôpitaux de Paris, Hôpital Européen Georges Pompidou, Service d'Hématologie Biologique, Paris, France;
authors: Pascale
authors: P
authors: Gaussem
mesh_terms: D000818:Animals; D001692:Biological Transport; D001792:Blood Platelets / Q000378:metabolism / Q000473:pathology*; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D006706:Homeostasis / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000502:physiology*; D010766:Phosphorylation; D015539:Platelet Activation; D010974:Platelet Aggregation; D010979:Platelet Function Tests; D015398:Signal Transduction; D013927:Thrombosis / Q000378:metabolism / Q000473:pathology*
chemical_list: D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D027425:ATP-Binding Cassette, Sub-Family C Proteins; C489540:Abcc4 protein, mouse