title: Monomeric C-reactive protein is prothrombotic and dissociates from circulating pentameric C-reactive protein on adhered activated platelets under flow.
abstract: OBJECTIVE
  We previously reported that C-reactive protein bioactivity on thrombogenesis was based on loss of its pentameric symmetry, resulting in formation of monomeric C-reactive protein. Our purpose was to provide mechanistic information on the direct effects of C-reactive protein isoforms on platelet activation and provide a C-reactive protein dissociation mechanism in circulating blood.
  
  
  RESULTS
  C-reactive protein-induced platelet activation was evaluated by flow cytometry. Platelet aggregation, clot properties, and coagulation were also measured. Washed platelets were incubated with C-reactive protein isoforms and vasodilator-stimulated phosphoprotein (VASP) phosphorylation was analysed by western blot and immunofluorescence. C-reactive protein dissociation under flow was evaluated by confocal microscopy on the surface of adhered platelets after perfusing human blood containing pentameric C-reactive protein at different shear rates. Dissociated monomeric C-reactive protein thrombogenicity was measured in flow experiments. Platelet aggregation and flow cytometry analysis revealed that monomeric C-reactive protein significantly induced platelet aggregation, surface P-selectin and CD63 exposure, and glycoprotein IIb-IIIa activation, whereas pentameric C-reactive protein was unable to produce any effect. p38 mitogen-activated protein kinase (MAPK) and Jun N-terminal kinase (JNK) inhibitors, as well as CD36 blocking antibody partially inhibited monomeric C-reactive protein-induced platelet activation and aggregation. Additionally, monomeric C-reactive protein significantly induced VASP dephosphorylation at serine 239. We found that pentameric C-reactive protein dissociated into monomeric C-reactive protein on the surface of activated adhered platelets under flow conditions and that this generated monomeric C-reactive protein promoted further platelet recruitment.
  
  
  CONCLUSIONS
  These data indicate that whereas serum pentameric C-reactive protein may not affect platelet activation, monomeric C-reactive protein, which dissociates from pentameric C-reactive protein on the surface of activated platelets, could contribute to atherothrombotic complications by promoting thrombosis.
authors.affiliation: Cardiovascular Research Center, CSIC-ICCC,  Institut Investigacions Biomèdiques Sant Pau, Hospital de la Santa Creu i Sant Pau, C/ Sant Antoni Mª Claret 167, 08025 Barcelona,  Spain.
authors.forename: Blanca
authors.initials: B
authors.lastname: Molins
authors.forename: Esther
authors.initials: E
authors.lastname: Peña
authors.forename: Raquel
authors.initials: R
authors.lastname: de la Torre
authors.forename: Lina
authors.initials: L
authors.lastname: Badimon
mesh_terms: D001777:Blood Coagulation; D001792:Blood Platelets / Q000378:metabolism*; D015153:Blotting, Western; D002097:C-Reactive Protein / Q000737:chemistry / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000097:blood; D006152:Cyclic GMP / Q000097:blood; D017869:Cyclic GMP-Dependent Protein Kinases / Q000097:blood; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D008840:Microfilament Proteins / Q000097:blood; D018613:Microscopy, Confocal; D010750:Phosphoproteins / Q000097:blood; D010766:Phosphorylation; D010973:Platelet Adhesiveness; D010974:Platelet Aggregation; D010979:Platelet Function Tests; D055503:Protein Multimerization; D012039:Regional Blood Flow; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D013927:Thrombosis / Q000097:blood*; D013997:Time Factors; D000098662:Vasodilator-Stimulated Phosphoprotein
chemical_list: D015815:Cell Adhesion Molecules; D008840:Microfilament Proteins; D010750:Phosphoproteins; D000098662:Vasodilator-Stimulated Phosphoprotein; D002097:C-Reactive Protein; D017869:Cyclic GMP-Dependent Protein Kinases; D006152:Cyclic GMP