title: Peroxynitrite--altered platelet mitochondria--a new link between inflammation and hemostasis.
abstract: Using porcine blood, we tested the hypothesis that peroxynitrite (ONOO(-)) may affect platelet-fibrin clot formation, clot retraction rate (CRR) and fibrinolysis through the inhibition of platelet energy production. It was found that ONOO(-) reduces CRR and enlarges final clot size in platelet rich plasma (PRP) (IC(50)=100μM) and in whole blood (IC(50)=200μM) dose-dependently. In a reconstituted system (washed platelets+fibrinogen), CRR was inhibited by 5-100nM ONOO(-) (IC(50)=25nM). Concentrations of ONOO(-) reducing CRR in PRP, inhibited platelet oxygen consumption, augmented lactate production and decreased total ATP contents in clots derived from PRP. In washed platelets ONOO(-) (5-20nM) produced a drop of the mitochondrial transmembrane potential (ΔΨ(m)). Blocking of mitochondrial energy production resulted in a reduction of CRR, whereas inhibition of glycolysis failed to affect CRR. ONOO(-), up to 300μM, failed to affect coagulation in platelet free plasma. Fibrinolysis of platelet-fibrin clots was enhanced by ONOO(-) (25-300μM), cytochalasin B and following the reduction of platelet energy production. Fibrinolysis of plasma clots was resistant to ONOO(-) treatment up to a concentration of 500μM. Tromboelastometry (ROTEM) measurements performed in PRP show that inhibition of platelet energy production or treatment with ONOO(-) (100-300μM) diminishes MCF, alpha angle and MCE parameters. Blockage the platelet contractile apparatus by cytochalasin B resulted in reduction of CRR and ROTEM variables (MCF, alpha angle, MCE). We conclude that physiologically relevant ONOO(-) concentrations may inhibit clot retraction, reduce clot stability and accelerate its lysis through the inhibition of platelet mitochondrial energy production.
authors.affiliation: Department of Physical Chemistry, Medical University of Bialystok, Kilinskiego 1, 15-089 Bialystok, Poland.
authors.forename: Tomasz
authors.initials: T
authors.lastname: Misztal
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Przesław
authors.forename: Tomasz
authors.initials: T
authors.lastname: Rusak
authors.forename: Marian
authors.initials: M
authors.lastname: Tomasiak
mesh_terms: D000199:Actins / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000097:blood; D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D004734:Energy Metabolism / Q000187:drug effects; D005260:Female; D005342:Fibrinolysis / Q000187:drug effects; D007249:Inflammation / Q000097:blood* / Q000276:immunology; D007700:Kinetics; D019344:Lactic Acid / Q000097:blood; D008297:Male; D053078:Membrane Potential, Mitochondrial; D008928:Mitochondria / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D010101:Oxygen Consumption; D030421:Peroxynitrous Acid / Q000097:blood*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D058105:Polymerization; D034421:Sus scrofa; D013916:Thrombelastography
chemical_list: D000199:Actins; D010975:Platelet Aggregation Inhibitors; D030421:Peroxynitrous Acid; D019344:Lactic Acid; D000255:Adenosine Triphosphate