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Thrombin flux and wall shear rate regulate fibrin fiber deposition state during polymerization under flow.

ABSTRACT: Thrombin is released as a soluble enzyme from the surface of platelets and tissue-factor-bearing cells to trigger fibrin polymerization during thrombosis under flow conditions. Although isotropic fibrin polymerization under static conditions involves protofibril extension and lateral aggregation leading to a gel, factors regulating fiber growth are poorly quantified under hemodynamic flow due to the difficulty of setting thrombin fluxes. A membrane microfluidic device allowed combined control of both thrombin wall flux (10(-13) to 10(-11) nmol/mum(2) s) and the wall shear rate (10-100 s(-1)) of a flowing fibrinogen solution. At a thrombin flux of 10(-12) nmol/mum(2) s, both fibrin deposition and fiber thickness decreased as the wall shear rate increased from 10 to 100 s(-1). Direct measurement and transport-reaction simulations at 12 different thrombin flux-wall shear rate conditions demonstrated that two dimensionless numbers, the Peclet number (Pe) and the Damkohler number (Da), defined a state diagram to predict fibrin morphology. For Da < 10, we only observed thin films at all Pe. For 10 < Da < 900, we observed either mat fibers or gels, depending on the Pe. For Da > 900 and Pe < 100, we observed three-dimensional gels. These results indicate that increases in wall shear rate quench first lateral aggregation and then protofibril extension.

SUBMITTER: Neeves KB

PROVIDER: S-EPMC2849060 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Thrombin flux and wall shear rate regulate fibrin fiber deposition state during polymerization under flow.

Neeves K B KB Illing D A R DA Diamond S L SL

Biophysical journal 20100401 7

Thrombin is released as a soluble enzyme from the surface of platelets and tissue-factor-bearing cells to trigger fibrin polymerization during thrombosis under flow conditions. Although isotropic fibrin polymerization under static conditions involves protofibril extension and lateral aggregation leading to a gel, factors regulating fiber growth are poorly quantified under hemodynamic flow due to the difficulty of setting thrombin fluxes. A membrane microfluidic device allowed combined control of ...[more]

PMID: 20371335

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Project description:Thrombin is an enzyme produced during blood coagulation that is crucial to the formation of a stable clot. Thrombin cleaves soluble fibrinogen into fibrin, which polymerizes and forms an insoluble, stabilizing gel around the growing clot. A small fraction of circulating fibrinogen is the variant γA/γ′, which has been associated with high-affinity thrombin binding and implicated as a risk factor for myocardial infarctions, deep vein thrombosis, and coronary artery disease. Thrombin is also known to be strongly sequestered by polymerized fibrin for extended periods of time in a way that is partially regulated by γA/γ′. However, the role of γA/γ′-thrombin interactions during fibrin polymerization is not fully understood. Here, we present a mathematical model of fibrin polymerization that considered the interactions between thrombin, fibrinogen, and fibrin, including those with γA/γ′. In our model, bivalent thrombin-fibrin binding greatly increased thrombin residency times and allowed for thrombin-trapping during fibrin polymerization. Results from the model showed that early in fibrin polymerization, γ′ binding to thrombin served to localize the thrombin to the fibrin(ogen), which effectively enhanced the enzymatic conversion of fibrinogen to fibrin. When all the fibrin was fully generated, however, the fibrin-thrombin binding persisted but the effect of fibrin on thrombin switched quickly to serve as a sink, essentially removing all free thrombin from the system. This dual role for γ′-thrombin binding during polymerization led to a paradoxical decrease in trapped thrombin as the amount of γ′ was increased. The model highlighted biochemical and biophysical roles for fibrin-thrombin interactions during polymerization and agreed well with experimental observations. Author summary We developed a mathematical model of fibrin polymerization that explicitly incorporated thrombin-fibrin(ogen) interactions, including the γ′ variant of fibrin(ogen). This variant is associated with clots that are more resistant to fibrinolysis and is therefore implicated as a risk factor in cardiovascular disease. However, the underlying mechanism it is not completely understood. We previously modeled γ′-thrombin interactions in a preformed fibrin clot and hypothesized that some thrombin became physically trapped within the clot during its formation (fibrin polymerization). We developed the model in this study to test that hypothesis and determine the role of γ′ during polymerization. The new model showed the plausibility of large amounts of thrombin being trapped within fibrin fibers during polymerization and suggested a dual role for γ′ during polymerization: localization of thrombin to the fibrin(ogen) during an initial early phase of polymerization and sequestration of thrombin during the later phase. The model results suggested a new regulatory mechanism for fibrin polymerization.

Project description:BackgroundCotton fiber is a single cell that arises from the epidermis of ovule. It is not only a main economic product of cotton, but an ideal material for studying on the growth and development of plant cell. Our previous study indicated that phytosterol content and the ratio of campesterol to sitosterol fluctuated regularly in cotton fiber development. However, what effects of modified phytosterol content and composition on the growth and development of cotton fiber cell is unknown. In this study, we overexpressed the GhSMT2-1, a cotton homologue of sterol C-24 methyltransferase 2 gene in transgenic upland cotton plants to modify phytosterol content and composition in fiber cells and investigated the changes on fiber elongation and secondary cell wall deposition.ResultsGhSMT2-1 overexpression led to changes of phytosterol content and the ratio of campesterol to sitosterol in fiber cell. At the rapid elongation stage of fiber cell, total phytosterol and sitosterol contents were increased while campesterol content was decreased in transgenic fibers when compared to control fibers. Accordingly, the ratio of campesterol to sitosterol declined strikingly. Simultaneously, the transgenic fibers were shorter and thicker than control fibers. Exogenous application of sitosterol or campesterol separately inhibited control fiber cell elongation in cotton ovule culture system in vitro. In addition, campesterol treatment partially rescued transgenic fiber elongation.ConclusionThese results elucidated that modification of phytosterol content and composition influenced fiber cell elongation and secondary cell wall formation. High sitosterol or low ratio of campesterol to sitosterol suppresses fiber elongation and/or promote secondary cell wall deposition. The roles of sitosterol and campesterol were discussed in fiber cell development. There might be a specific ratio of campesterol to sitosterol in different developmental stage of cotton fibers, in which GhSMT2-1 play an important role. Our study, at a certain degree, provides novel insights into the regulatory mechanisms of fiber cell development.

Project description:ObjectiveAt sites of vascular injury, thrombin is an important mediator in thrombus growth and stability. Using microfluidic flow devices as well as patterned surfaces of collagen and tissue factor (TF), we sought to determine the role that fibrin plays in clot stability without interfering with the production of thrombin.Methods and resultsWe deployed an 8-channel microfluidic device to study coagulation during corn trypsin inhibitor-treated (XIIa-inhibited) whole blood perfusion over lipidated TF linked to a fibrillar collagen type 1 surface. Clot growth and embolization were measured at initial inlet venous (200 s(-1)) or arterial (1000 s(-1)) wall shear rates under constant flow rate or pressure relief mode in the presence or absence of Gly-Pro-Arg-Pro (GPRP) to block fibrin polymerization. Numerical calculations for each mode defined hemodynamic forces on the growing thrombi. In either mode at inlet venous flow, increasing amounts of TF on the surface led to a modest dose-dependent increase (up to 2-fold) in platelet deposition, but resulted in massive fibrin accumulation (>50-fold) only when exceeding a critical TF threshold. At a venous inlet flow, GPRP led to a slight 20% increase in platelet accumulation (P<0.01) in pressure relief mode with thrombi resisting ≈1500 s(-1) before full channel occlusion. GPRP-treated thrombi were unstable under constant flow rate, where shear forces caused embolization at a maximum shear rate of ≈2300 s(-1) (69 dynes/cm2). In constant flow rate mode, the nonocclusive platelet-fibrin deposits (no GPRP) withstood maximum shear rates of ≈29 000 s(-1) (870 dyne/cm2) at ≈95% of full channel occlusion. For arterial inlet shear rate, embolization was marked for either mode with GPRP present when shear forces reached 87 dynes/cm2 (≈2900 s(-1)). Under constant flow rate, platelet-fibrin deposits (no GPRP) withstood maximums of 2400 dynes/cm2 (80,000 s(-1)) at ≈90% of full channel occlusion prior to embolization.ConclusionsFibrin increased clot strength by 12- to 28-fold. Under pressure relief mode, ≈2-fold more fibrin was produced under venous flow (P<0.001). These studies define embolization criteria for clots formed with surface TF-triggered thrombin production (±fibrin) under venous and arterial flows.

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Thrombin flux and wall shear rate regulate fibrin fiber deposition state during polymerization under flow.

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Thrombin flux and wall shear rate regulate fibrin fiber deposition state during polymerization under flow.

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