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Physical determinants of fibrinolysis in single fibrin fibers.

Bucay I, O'Brien ET, Wulfe SD, Superfine R, Wolberg AS, Falvo MR, Hudson NE - PLoS ONE (2015)

Bottom Line: We found that during lysis 64 ± 6% of fibers were transected at one point, but 29 ± 3% of fibers increase in length rather than dissolving or being transected.Because lysis rates were greatly reduced in elongated fibers, we hypothesize that plasmin activity depends on fiber strain.These results highlight how subtle differences in the diameter and prestrain of fibers could lead to dramatically different lytic susceptibilities.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics and Astronomy, University of North Carolina, Chapel Hill, North Carolina, United States of America.

ABSTRACT
Fibrin fibers form the structural backbone of blood clots; fibrinolysis is the process in which plasmin digests fibrin fibers, effectively regulating the size and duration of a clot. To understand blood clot dissolution, the influence of clot structure and fiber properties must be separated from the effects of enzyme kinetics and perfusion rates into clots. Using an inverted optical microscope and fluorescently-labeled fibers suspended between micropatterned ridges, we have directly measured the lysis of individual fibrin fibers. We found that during lysis 64 ± 6% of fibers were transected at one point, but 29 ± 3% of fibers increase in length rather than dissolving or being transected. Thrombin and plasmin dose-response experiments showed that the elongation behavior was independent of plasmin concentration, but was instead dependent on the concentration of thrombin used during fiber polymerization, which correlated inversely with fiber diameter. Thinner fibers were more likely to lyse, while fibers greater than 200 ± 30 nm in diameter were more likely to elongate. Because lysis rates were greatly reduced in elongated fibers, we hypothesize that plasmin activity depends on fiber strain. Using polymer physics- and continuum mechanics-based mathematical models, we show that fibers polymerize in a strained state and that thicker fibers lose their prestrain more rapidly than thinner fibers during lysis, which may explain why thick fibers elongate and thin fibers lyse. These results highlight how subtle differences in the diameter and prestrain of fibers could lead to dramatically different lytic susceptibilities.

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Modeling the lysis of a Fiber Consisting of an Inner Core and Outer Shell.A Left: cross-section of a fiber with an outer shell (A) and inner core (B). Right: a portrayal of the relative lengths of the shell, the core, the free fiber (equilibrium fiber length), and SS-suspended fiber. B The fiber free length, normalized to the length between the structured surfaces, for thin (blue, 40 nm radius), medium (red, 80 nm radius), and thick (green, 120 nm radius) fibers as the shell is lysed. The black line signifies the SS length (LSS) and the black dot is the free length of the fiber after 100% lysis of the shell (i.e., the core length). Note that there is no radial dependence on the free length for fibers of the constant ratio model. For this plot LoA = 18 μm, S = 1.2. For the constant core, the core thickness was 15 nm.
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pone.0116350.g005: Modeling the lysis of a Fiber Consisting of an Inner Core and Outer Shell.A Left: cross-section of a fiber with an outer shell (A) and inner core (B). Right: a portrayal of the relative lengths of the shell, the core, the free fiber (equilibrium fiber length), and SS-suspended fiber. B The fiber free length, normalized to the length between the structured surfaces, for thin (blue, 40 nm radius), medium (red, 80 nm radius), and thick (green, 120 nm radius) fibers as the shell is lysed. The black line signifies the SS length (LSS) and the black dot is the free length of the fiber after 100% lysis of the shell (i.e., the core length). Note that there is no radial dependence on the free length for fibers of the constant ratio model. For this plot LoA = 18 μm, S = 1.2. For the constant core, the core thickness was 15 nm.

Mentions: Fibrin fibers have a well-defined 23 nm banding pattern when viewed by electron microscopy [36]. The banding has been ascribed to protofibrils wrapping outwardly around the center of the fiber and in registry with the inner protofibrils; for this to be true, the outer protofibrils must be stretched to align with the inner molecules [21,27]. Based on this picture, we propose a simplified core-shell model with two mechanical “domains” with the same elastic modulus E: domain A (outer shell, which is stretched) and domain B (inner core, which is compressed) (Fig. 5A). In a taut fiber suspended across a structured surface (SS) or within a fibrin clot network, the shell is stretched and the core is compressed. The free fiber length LF is the equilibrium length of the conjoined domains and the requirement that a fiber be taut pre-fibrinolysis necessitates the condition that LF < LSS. LF, will therefore change as the outer shell is lysed as given by:LF'=SLoA((1−x)R2+xrB2S(1−x)(R2−rB2)+rB2)Where R is the fiber radius, rB is the core radius, x is the percentage of the outer shell lysed, and we have imposed the constraint that LoB = SLoA, where S > 1 is a proportionality constant relating LoB and LoA, and LoB and LoA are the initial length of the core and shell respectively (Fig. 5A). In this model, if fibrin fibers of all diameters share a common outer shell thickness, then thicker fibers elongate at a much earlier stage of fibrinolysis than thinner fibers, in agreement with our experimental data (Fig. 5B). Two other models of fibrin architecture (constant core radius, and constant core:shell radius ratio) do not show the correct increase in length as the fiber lyses (see S1 File and S2 Fig.). Together, our experimental and modeling data indicate that fiber diameter determines the prestrain induced by fibrin polymerization, and the pre-strain, in part, governs a fiber’s susceptibility to plasmin. Combining our models involving fiber strain during lysis, with other recently published models of lysis involving stochastic reactions and enzyme diffusion may be required for a complete picture of fibrinolysis [37,38].


Physical determinants of fibrinolysis in single fibrin fibers.

Bucay I, O'Brien ET, Wulfe SD, Superfine R, Wolberg AS, Falvo MR, Hudson NE - PLoS ONE (2015)

Modeling the lysis of a Fiber Consisting of an Inner Core and Outer Shell.A Left: cross-section of a fiber with an outer shell (A) and inner core (B). Right: a portrayal of the relative lengths of the shell, the core, the free fiber (equilibrium fiber length), and SS-suspended fiber. B The fiber free length, normalized to the length between the structured surfaces, for thin (blue, 40 nm radius), medium (red, 80 nm radius), and thick (green, 120 nm radius) fibers as the shell is lysed. The black line signifies the SS length (LSS) and the black dot is the free length of the fiber after 100% lysis of the shell (i.e., the core length). Note that there is no radial dependence on the free length for fibers of the constant ratio model. For this plot LoA = 18 μm, S = 1.2. For the constant core, the core thickness was 15 nm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4340865&req=5

pone.0116350.g005: Modeling the lysis of a Fiber Consisting of an Inner Core and Outer Shell.A Left: cross-section of a fiber with an outer shell (A) and inner core (B). Right: a portrayal of the relative lengths of the shell, the core, the free fiber (equilibrium fiber length), and SS-suspended fiber. B The fiber free length, normalized to the length between the structured surfaces, for thin (blue, 40 nm radius), medium (red, 80 nm radius), and thick (green, 120 nm radius) fibers as the shell is lysed. The black line signifies the SS length (LSS) and the black dot is the free length of the fiber after 100% lysis of the shell (i.e., the core length). Note that there is no radial dependence on the free length for fibers of the constant ratio model. For this plot LoA = 18 μm, S = 1.2. For the constant core, the core thickness was 15 nm.
Mentions: Fibrin fibers have a well-defined 23 nm banding pattern when viewed by electron microscopy [36]. The banding has been ascribed to protofibrils wrapping outwardly around the center of the fiber and in registry with the inner protofibrils; for this to be true, the outer protofibrils must be stretched to align with the inner molecules [21,27]. Based on this picture, we propose a simplified core-shell model with two mechanical “domains” with the same elastic modulus E: domain A (outer shell, which is stretched) and domain B (inner core, which is compressed) (Fig. 5A). In a taut fiber suspended across a structured surface (SS) or within a fibrin clot network, the shell is stretched and the core is compressed. The free fiber length LF is the equilibrium length of the conjoined domains and the requirement that a fiber be taut pre-fibrinolysis necessitates the condition that LF < LSS. LF, will therefore change as the outer shell is lysed as given by:LF'=SLoA((1−x)R2+xrB2S(1−x)(R2−rB2)+rB2)Where R is the fiber radius, rB is the core radius, x is the percentage of the outer shell lysed, and we have imposed the constraint that LoB = SLoA, where S > 1 is a proportionality constant relating LoB and LoA, and LoB and LoA are the initial length of the core and shell respectively (Fig. 5A). In this model, if fibrin fibers of all diameters share a common outer shell thickness, then thicker fibers elongate at a much earlier stage of fibrinolysis than thinner fibers, in agreement with our experimental data (Fig. 5B). Two other models of fibrin architecture (constant core radius, and constant core:shell radius ratio) do not show the correct increase in length as the fiber lyses (see S1 File and S2 Fig.). Together, our experimental and modeling data indicate that fiber diameter determines the prestrain induced by fibrin polymerization, and the pre-strain, in part, governs a fiber’s susceptibility to plasmin. Combining our models involving fiber strain during lysis, with other recently published models of lysis involving stochastic reactions and enzyme diffusion may be required for a complete picture of fibrinolysis [37,38].

Bottom Line: We found that during lysis 64 ± 6% of fibers were transected at one point, but 29 ± 3% of fibers increase in length rather than dissolving or being transected.Because lysis rates were greatly reduced in elongated fibers, we hypothesize that plasmin activity depends on fiber strain.These results highlight how subtle differences in the diameter and prestrain of fibers could lead to dramatically different lytic susceptibilities.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics and Astronomy, University of North Carolina, Chapel Hill, North Carolina, United States of America.

ABSTRACT
Fibrin fibers form the structural backbone of blood clots; fibrinolysis is the process in which plasmin digests fibrin fibers, effectively regulating the size and duration of a clot. To understand blood clot dissolution, the influence of clot structure and fiber properties must be separated from the effects of enzyme kinetics and perfusion rates into clots. Using an inverted optical microscope and fluorescently-labeled fibers suspended between micropatterned ridges, we have directly measured the lysis of individual fibrin fibers. We found that during lysis 64 ± 6% of fibers were transected at one point, but 29 ± 3% of fibers increase in length rather than dissolving or being transected. Thrombin and plasmin dose-response experiments showed that the elongation behavior was independent of plasmin concentration, but was instead dependent on the concentration of thrombin used during fiber polymerization, which correlated inversely with fiber diameter. Thinner fibers were more likely to lyse, while fibers greater than 200 ± 30 nm in diameter were more likely to elongate. Because lysis rates were greatly reduced in elongated fibers, we hypothesize that plasmin activity depends on fiber strain. Using polymer physics- and continuum mechanics-based mathematical models, we show that fibers polymerize in a strained state and that thicker fibers lose their prestrain more rapidly than thinner fibers during lysis, which may explain why thick fibers elongate and thin fibers lyse. These results highlight how subtle differences in the diameter and prestrain of fibers could lead to dramatically different lytic susceptibilities.

Show MeSH
Related in: MedlinePlus