Fibrin fibers form the structural backbone of blood clots. The structural properties of fibrin clots are highly dependent on formation kinetics. Environmental factors such as protein concentration, pH, salt, and protein modification, to name a few, can affect fiber kinetics through altered fibrinopeptide release, monomer association, and/or lateral aggregation. The objective of our study was to determine the effect of thrombin and fibrinogen exposed to nitric oxide on fibrin clot properties. ProliNONOate (5 [mu]mol/l) was added to fibrinogen and thrombin before clot initiation and immediately following the addition of thrombin to the fibrinogen solution. Resulting fibrin fibers were probed with an atomic force microscope to determine their diameter and extensibility and fibrin clots were analyzed for clot density using confocal microscopy. Fiber diameters were also determined by confocal microscopy and the rate of clot formation was recorded using UV-vis spectrophotometry. Protein oxidation and S-nitrosation was determined by UV-vis, ELISA, and chemiluminescence. The addition of ProliNONOate to fibrinogen or thrombin resulted in a change in clot structure. ProliNONOate exposure produced clots with lower fiber density, thicker fibers, and increased time to maximum turbidity. The effect of the exposure of nitric oxide to thrombin and fibrinogen were measured independently and indicated that each plays a role in altering clot properties. We detected thrombin S-nitrosation and protein carbonyl formation after nitric oxide exposure. Our study reveals a regulation of fibrin clot properties by nitric oxide exposure and suggests a role of peroxynitrite in oxidative modifications of the proteins. These results relate nitric oxide bioavailability and oxidative stress to altered clot properties.

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Copyright © 2016 Wolters Kluwer Health, Inc. Article first published online: 13 Oct 2016.

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DOI: 10.1097/MBC.0000000000000602

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Helms, Christine C., Shannon Kapadia, Anne C. Gilmore, Zhexi Lu, Swati Basu, and Daniel B. Kim-Shapiro. "Exposure of fibrinogen and thrombin to nitric oxide donor ProliNONOate affects fibrin clot properties." Blood Coagulation & Fibrinolysis: An International Journal in Haemostasis and Thrombosis (October 13, 2016). doi: 10.1097/MBC.0000000000000602.