title: Redox-based thrombelastographic method to detect carboxyhemefibrinogen-mediated hypercoagulability.
abstract: Cigarette smoking is associated with plasmatic hypercoagulability, and carbon monoxide has been demonstrated to enhance coagulation by binding to a fibrinogen-bound heme. Our objective was to design and test a redox-based method to detect carboxyhemefibrinogen. Normal, pooled, citrated plasma was exposed to 0-100  μmol/l carbon monoxide releasing molecule-2 (tricarbonyldichlororuthenium (II) dimer; CORM-2) before or after exposure to the organic reductant phenylhydroxylamine (PHA, 0-30  mmol/l), a compound that rapidly converts Fe(+2) to Fe(+3) in heme. Addition of calcium and tissue factor activation in disposable thrombelastographic cups was performed, followed by data collection at 37°C for 15  min. Elastic modulus (G, dynes/cm(2)) was the primary endpoint. CORM-2 significantly increased G values by 67.8% compared to unexposed plasma; pretreatment with 10  mmol/l PHA significantly decreased G values in CORM-2-exposed plasma by 77.1%, whereas 30  mmol/l PHA was required to significantly decrease G values by 64.0% in plasma following CORM-2 pre-exposure. G values were not significantly different between unexposed plasma and plasma exposed to CORM-2 followed by 30  mmol/l PHA addition. Conversion of fibrinogen-bound to the metheme state alone decreased G by 34.3-38.9% following exposure to 10-30  mmol/l PHA. Conversion of fibrinogen-bound heme Fe(+2) to Fe(+3) with PHA abrogated carbon monoxide-mediated increases in clot strength. Clinical trials are planned to investigate smoking individuals to mechanistically link carboxyhemefibrinogen formation with in-vitro hypercoagulability.
authors.affiliation: Department of Anesthesiology, Drexel University College of Medicine, Philadelphia, Pennsylvania, USA. vance.nielsen@drexelmed.edu
authors.forename: Vance G
authors.initials: VG
authors.lastname: Nielsen
authors.forename: Matthew R
authors.initials: MR
authors.lastname: Arkebauer
authors.forename: Keith
authors.initials: K
authors.lastname: Vosseller
mesh_terms: D001777:Blood Coagulation / Q000187:drug effects*; D002118:Calcium / Q000494:pharmacology; D002248:Carbon Monoxide / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D055119:Elastic Modulus / Q000187:drug effects*; D005340:Fibrinogen / Q000378:metabolism; D006418:Heme / Q000378:metabolism; D006801:Humans; D006898:Hydroxylamines / Q000494:pharmacology; D009942:Organometallic Compounds / Q000009:adverse effects; D010084:Oxidation-Reduction; D010949:Plasma / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D012907:Smoking / Q000009:adverse effects / Q000097:blood*; D013916:Thrombelastography / Q000379:methods*; D019851:Thrombophilia / Q000097:blood*; D013925:Thromboplastin / Q000032:analysis / Q000096:biosynthesis
chemical_list: D006898:Hydroxylamines; D009942:Organometallic Compounds; C447082:tricarbonyldichlororuthenium (II) dimer; C012484:phenylhydroxylamine; D006418:Heme; D002248:Carbon Monoxide; D005340:Fibrinogen; D013925:Thromboplastin; D002118:Calcium