OBJECTIVE: Peroxynitrite (ONOO-) is an oxidant formed from the rapid reaction of superoxide and nitric oxide (NO) at sites of inflammation. The literature reports conflicting data on the effects of ONOO- in biological systems, with both NO- and oxidant-dependent effects having been demonstrated. The aim of this study was to investigate these distinct mechanisms through examining molecular aspects of the effects of ONOO- on human platelets, a system in which we have previously shown that ONOO- has both pro- and anti-aggregatory effects. METHODS: Platelet function was assessed by measuring platelet P-selectin expression flow cytometrically, intraplatelet Ca2+ concentrations, and by light aggregometry. A colorimetric method was used to measure extracellular platelet membrane thiols. The contribution of NO and cGMP to the pharmacological effects of ONOO- was investigated using an inhibitor of the soluble guanylate cyclase (sGC), 1H-[1,2,4]oxadiazolo[4,3-alpha]quinoxalin-1-one (ODQ), and the NO scavenger oxy-haemoglobin. RESULTS: Peroxynitrite (50-400 microM) caused a concentration-dependent increase in the number of platelets expressing P-selectin, an increase in intraplatelet Ca2+ concentrations and a decrease in platelet membrane thiols. Peroxynitrite-induced P-selectin expression was augmented by ODQ. In contrast, when P-selectin expression was elicited by collagen, ONOO- acted as an inhibitor of this process, an effect that was further enhanced by the addition of 1% plasma, ODQ or oxy-haemoglobin abolished this inhibitory effect. Finally, low concentrations (50-100 microM) of ONOO- inhibited collagen-induced platelet aggregation, an effect that was reversed by oxy-haemoglobin. CONCLUSIONS: Peroxynitrite exerts dual effects on platelets, which are either activating or inhibitory due to the conversion of ONOO- to NO or NO donors. Peroxynitrite-induced platelet activation seems to be due to thiol oxidation and an increase in intracellular Ca2+. It is important to note that inhibitory, NO-dependent effects occur at lower concentrations than the activating effects. These data are then consistent with the conflicting literature, showing both damaging and cytoprotective effects of ONOO- in biological systems. We hypothesize that the conversion of ONOO- to NO is the critical factor determining the outcome of ONOO- exposure in vivo.
OBJECTIVE:Peroxynitrite (ONOO-) is an oxidant formed from the rapid reaction of superoxide and nitric oxide (NO) at sites of inflammation. The literature reports conflicting data on the effects of ONOO- in biological systems, with both NO- and oxidant-dependent effects having been demonstrated. The aim of this study was to investigate these distinct mechanisms through examining molecular aspects of the effects of ONOO- on human platelets, a system in which we have previously shown that ONOO- has both pro- and anti-aggregatory effects. METHODS: Platelet function was assessed by measuring platelet P-selectin expression flow cytometrically, intraplatelet Ca2+ concentrations, and by light aggregometry. A colorimetric method was used to measure extracellular platelet membrane thiols. The contribution of NO and cGMP to the pharmacological effects of ONOO- was investigated using an inhibitor of the soluble guanylate cyclase (sGC), 1H-[1,2,4]oxadiazolo[4,3-alpha]quinoxalin-1-one (ODQ), and the NO scavenger oxy-haemoglobin. RESULTS:Peroxynitrite (50-400 microM) caused a concentration-dependent increase in the number of platelets expressing P-selectin, an increase in intraplatelet Ca2+ concentrations and a decrease in platelet membrane thiols. Peroxynitrite-induced P-selectin expression was augmented by ODQ. In contrast, when P-selectin expression was elicited by collagen, ONOO- acted as an inhibitor of this process, an effect that was further enhanced by the addition of 1% plasma, ODQ or oxy-haemoglobin abolished this inhibitory effect. Finally, low concentrations (50-100 microM) of ONOO- inhibited collagen-induced platelet aggregation, an effect that was reversed by oxy-haemoglobin. CONCLUSIONS:Peroxynitrite exerts dual effects on platelets, which are either activating or inhibitory due to the conversion of ONOO- to NO or NO donors. Peroxynitrite-induced platelet activation seems to be due to thiol oxidation and an increase in intracellular Ca2+. It is important to note that inhibitory, NO-dependent effects occur at lower concentrations than the activating effects. These data are then consistent with the conflicting literature, showing both damaging and cytoprotective effects of ONOO- in biological systems. We hypothesize that the conversion of ONOO- to NO is the critical factor determining the outcome of ONOO- exposure in vivo.
Authors: Stephen R Thom; Veena M Bhopale; Shih-Tsung Han; James M Clark; Kevin R Hardy Journal: Am J Respir Crit Care Med Date: 2006-08-24 Impact factor: 21.405
Authors: Sisi Marcondes; Marcia H M Cardoso; Rafael P Morganti; Sara M Thomazzi; Sergio Lilla; Ferid Murad; Gilberto De Nucci; Edson Antunes Journal: Proc Natl Acad Sci U S A Date: 2006-02-21 Impact factor: 11.205
Authors: Jonathan Watt; Marie-Ann Ewart; Fiona H Greig; Keith G Oldroyd; Roger M Wadsworth; Simon Kennedy Journal: Thromb Res Date: 2012-04-19 Impact factor: 3.944