title: Extracellular histones increase plasma thrombin generation by impairing thrombomodulin-dependent protein C activation.
abstract: BACKGROUND
  Histones are basic proteins that contribute to cell injury and tissue damage when released into the extracellular space. They have been attributed a prothrombotic activity, because their injection into mice induces diffuse microvascular thrombosis. The protein C-thrombomodulin (TM) system is a fundamental regulator of coagulation, particularly in the microvasculature, and its activity can be differentially influenced by interaction with several cationic proteins.
  
  
  OBJECTIVE
  To evaluate the effect of histones on the protein C-TM system in a plasma thrombin generation assay and in purified systems.
  
  
  METHODS
  The effect of histones on plasma thrombin generation in the presence or absence of TM was analyzed by calibrated automated thrombinography. Protein C activation in purified systems was evaluated by chromogenic substrate cleavage. The binding of TM and protein C to histones was evaluated by solid-phase binding assay.
  
  
  RESULTS
  Histones dose-dependently increased plasma thrombin generation in the presence of TM, independently of its chondroitin sulfate moiety. This effect was not caused by inhibition of activated protein C activity, but by the impairment of TM-mediated protein C activation. Histones were able to bind to both protein C and TM, but the carboxyglutamic acid domain of protein C was required for their effect. Histones H4 and H3 displayed the highest activity. Importantly, unlike heparin, DNA did not inhibit the potentiating effect of histones on thrombin generation.
  
  
  CONCLUSIONS
  Histones enhance plasma thrombin generation by reducing TM-dependent protein C activation. This mechanism might contribute to microvascular thrombosis induced by histones in vivo at sites of organ failure or severe inflammation.
authors.affiliation: Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA.
authors.forename: C T
authors.initials: CT
authors.lastname: Ammollo
authors.forename: F
authors.initials: F
authors.lastname: Semeraro
authors.forename: J
authors.initials: J
authors.lastname: Xu
authors.forename: N L
authors.initials: NL
authors.lastname: Esmon
authors.forename: C T
authors.initials: CT
authors.lastname: Esmon
mesh_terms: D000818:Animals; D001777:Blood Coagulation / Q000187:drug effects / Q000502:physiology; D004247:DNA / Q000378:metabolism / Q000494:pharmacology; D005110:Extracellular Space / Q000378:metabolism; D006493:Heparin / Q000378:metabolism / Q000494:pharmacology; D006657:Histones / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D011485:Protein Binding; D011486:Protein C / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013917:Thrombin / Q000096:biosynthesis*; D018180:Thrombomodulin / Q000378:metabolism*; D013927:Thrombosis / Q000097:blood / Q000209:etiology
chemical_list: D006657:Histones; D011486:Protein C; D011994:Recombinant Proteins; C546931:THBD protein, human; D018180:Thrombomodulin; D006493:Heparin; D004247:DNA; D013917:Thrombin