Background 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. Objective To determine the effect of thrombin and fibrinogen exposed to nitric oxide on fibrin clot properties. Methods ProliNONOate (5 μM) 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. Results The addition of ProliNONOate to fibrinogen or thrombin resulted 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. Conclusions 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 NO bioavailability and oxidative stress to altered clot properties.

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