title: Platelet CD40L Modulates Thrombus Growth Via Phosphatidylinositol 3-Kinase β, and Not Via CD40 and IκB Kinase α.
abstract: OBJECTIVE
  To investigate the roles and signaling pathways of CD40L and CD40 in platelet-platelet interactions and thrombus formation under conditions relevant for atherothrombosis.
  
  
  RESULTS
  Platelets from mice prone to atherosclerosis lacking CD40L (Cd40lg(-/-)Apoe(-/-)) showed diminished αIIbβ3 activation and α-granule secretion in response to glycoprotein VI stimulation, whereas these responses of CD40-deficient platelets (Cd40(-/-)Apoe(-/-)) were not decreased. Using blood from Cd40lg(-/-)Apoe(-/-) and Cd40(-/-)Apoe(-/-) mice, the glycoprotein VI-dependent formation of dense thrombi was impaired on atherosclerotic plaque material or on collagen, in comparison with Apoe(-/-) blood. In all genotypes, addition of CD40L to the blood enhanced the growth of dense thrombi on plaques and collagen. Similarly, CD40L enhanced glycoprotein VI-induced platelet aggregation, even with platelets deficient in CD40. This potentiation was antagonized in Pik3cb(R/R) platelets or by inhibiting phosphatidylinositol 3-kinase β (PI3Kβ). Addition of CD40L also enhanced collagen-induced Akt phosphorylation, which was again antagonized by absence or inhibition of PI3Kβ. Finally, platelets from Chuk1(A/A)Apoe(-/-) mice deficient in IκB kinase α (IKKα), implicated in CD40 signaling to nuclear factor (NF) κB, showed unchanged responses to CD40L in aggregation or thrombus formation.
  
  
  CONCLUSIONS
  Under atherogenic conditions, CD40L enhances collagen-induced platelet-platelet interactions by supporting integrin αIIbβ3 activation, secretion and thrombus growth via PI3Kβ, but not via CD40 and IKKα/NFκB. This role of CD40L exceeds the no more than modest role of CD40 in thrombus formation.
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.). Marijke.Kuijpers@maastrichtuniversity.nl.
authors.forename: Marijke J E
authors.initials: MJ
authors.lastname: Kuijpers
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Nadine J A
authors.initials: NJ
authors.lastname: Mattheij
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Lina
authors.initials: L
authors.lastname: Cipolla
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Johanna P
authors.initials: JP
authors.lastname: van Geffen
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Toby
authors.initials: T
authors.lastname: Lawrence
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Marjo M P C
authors.initials: MM
authors.lastname: Donners
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Louis
authors.initials: L
authors.lastname: Boon
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Dirk
authors.initials: D
authors.lastname: Lievens
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Mauro
authors.initials: M
authors.lastname: Torti
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Heidi
authors.initials: H
authors.lastname: Noels
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Norbert
authors.initials: N
authors.lastname: Gerdes
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Judith M E M
authors.initials: JM
authors.lastname: Cosemans
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Esther
authors.initials: E
authors.lastname: Lutgens
authors.affiliation: From the Departments of Biochemistry (M.J.E.K., N.J.A.M., L.C., J.P.v.G., J.M.E.M.C., J.W.M.H.) and Molecular Genetics (M.M.P.C.D.), CARIM, Maastricht University, Maastricht, The Netherlands; Division of Biochemistry, Department of Biology and Biotechnology, University of Pavia, Pavia, Italy (L.C., M.T.); Centre d'Immunologie de Marseille-Luminy, Aix-Marseille Université, Marseille, France (T.L.); Bioceros, Utrecht, The Netherlands (L.B.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany (D.L., N.G., E.L.); Institute for Molecular Cardiovascular Research (IMCAR), RWTH Aachen University, Aachen, Germany (H.N.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (E.L.).
authors.forename: Johan W M
authors.initials: JW
authors.lastname: Heemskerk
mesh_terms: D000818:Animals; D050197:Atherosclerosis / Q000378:metabolism* / Q000473:pathology; D001792:Blood Platelets / Q000378:metabolism*; D019013:CD40 Antigens / Q000378:metabolism*; D023201:CD40 Ligand / Q000378:metabolism*; D003094:Collagen / Q000378:metabolism; D051550:I-kappa B Kinase / Q000378:metabolism*; D051379:Mice; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism*; D015539:Platelet Activation; D015398:Signal Transduction; D013927:Thrombosis / Q000378:metabolism* / Q000473:pathology
chemical_list: D019013:CD40 Antigens; D023201:CD40 Ligand; D003094:Collagen; D058539:Phosphatidylinositol 3-Kinase; D051550:I-kappa B Kinase
keywords: CD40; CD40 ligand; atherosclerosis; atherothrombosis; blood platelets; signal transduction; signaling pathways; thrombosis