Multiscale Mechanics of Fibrin Polymer: Gel Stretching with Protein Unfolding and Loss of Water

Abstract

Blood clots and thrombi consist primarily of a mesh of branched fibers made of the protein fibrin. We propose a molecular basis for the marked extensibility and negative compressibility of fibrin gels based on the structural and mechanical properties of clots at the network, fiber, and molecular levels. The force required to stretch a clot initially rises linearly and is accompanied by a dramatic decrease in clot volume and a peak in compressibility. These macroscopic transitions are accompanied by fiber alignment and bundling after forced protein unfolding. Constitutive models are developed to integrate observations at spatial scales that span six orders of magnitude and indicate that gel extensibility and expulsion of water are both manifestations of protein unfolding, which is not apparent in other matrix proteins such as collagen.Fibrin clots are proteinaceous gels that polymerize in the blood as a consequence of biochemical cascades at sites of vascular injury. Together with platelets, this meshwork stops bleeding and supports active contraction during wound healing (1,2). Fibrin also provides a scaffold for thrombi, clots that block blood vessels and cause tissue damage, leading to myocardial infarction, ischemic stroke, and other cardiovascular diseases (3). To maintain hemostasis while minimizing the impact of thrombosis, fibrin must have suitable stiffness and plasticity (4), but also sufficient permeability so that the network can be effectively decomposed (lysed) by proteolytic enzymes (5,6). It is challenging to meet all of these conditions because open scaffolds would be expected to break at low strains, as is true for collagen gels (7). To address how fibrin clots are both permeable and highly extensible, we studied fibrin structures across multiple spatial scales, from whole clots to single fibers and single molecules (Fig. 1).Fibrin clots were made from purified human fibrinogen under conditions (8) that resulted in the formation of long, straight fibers, similar to those found in physiological clots. To simplify the interpretation, the clots were covalently ligated with the use of a transglutaminase (blood clotting factor XIIIa), as naturally occurs in the blood, which prevents protofibrils from sliding past one another, thus eliminating persistent creep (9).Measurements of the extensibility of 2-mm-diameter fibrin clots ( Fig. 2A) demonstrated that the clots could be stretched to more than three times their relaxed length before breaking, with an average stretch of 2.7 ± 0.15-fold (n =6)(10). This is comparable to the single-fiber extensibility that is observed when a fibrin fiber is laterally stretched with an atomic force microscope (11). Qualitatively, the resulting force-strain curve for fibrin is similar to those observed for rubbers and other materials made from flexible chains (12). However, for fibrin clots, which are made of longer, straighter fibers than the thermally fluctuating polymer chains in rubber, models of rubber-like elasticity predict a branching density that is wro...

Differential Contributions of Conformation Extension and Domain Unfolding to Properties of Fibronectin Nanotextiles

“…12,25,28 Indeed, the force plateau observed at 80 pN is well captured using the two-state model, indicating that FN III domains unfold (solid line, Figure 3d). This observation Until now, the molecular mechanism accounting for FN elasticity has been debated between two structural models.…”

“…24 It was originally developed to describe the behavior of rubber and polymeric materials 24 but has recently been applied to understand the contribution of protein unfolding in fibrin protein gels. 25 The model idealizes a local, random network structure as semi-flexible chains connected at the center of a cube ( Figure 3c Figure 3d). However, if we assume that FN is in an extended conformation within the unloaded fabrics (r o = 160 nm 11,27 ), an extreme value of the force plateau (~8.6 nN) is predicted by the model (red axis, Figure 3d).…”

Interplay of Platelet Contractility and Elasticity of Fibrin/Erythrocytes in Blood Clot Retraction

“…This reorganization is driven by the energy-dependent interaction of nonmuscle myosin IIa with actin and is followed by the relocation of actin filaments to the filopodia on the periphery of the platelet (5). In addition to activating platelets, thrombin converts fibrinogen to fibrin (6)(7)(8)(9) and activates factor XIII (FXIII / FXIIIa), which catalyzes the cross-linking of fibrin (6). Cross-linking results in the stabilization of the viscoelastic fibrin network, which has many fibers originating from platelet aggregates (6)(7)(8)(9).…”

“…In addition to activating platelets, thrombin converts fibrinogen to fibrin (6)(7)(8)(9) and activates factor XIII (FXIII / FXIIIa), which catalyzes the cross-linking of fibrin (6). Cross-linking results in the stabilization of the viscoelastic fibrin network, which has many fibers originating from platelet aggregates (6)(7)(8)(9). The activated platelets are able to generate contractile forces that are propagated through the cross-linked fibrin fibers (10,11), effectively resulting in a reduction of the clot volume (12,13).…”

Fibrin structural and diffusional analysis suggests that fibers are permeable to solute transport

“…The storage modulus in the linear elastic regime is well-described by entropic models valid for semiflexible polymers [36], being governed by the total fiber length and the fiber persistence length, which in turn is dependent on the number of protofibrils per fiber. Dense fibrin networks with small fiber segment lengths between junctions and networks of thick fibers behave as athermal fibrous networks even at low strain [37,38].…”