A computational investigation of occlusive arterial thrombosis.

The generation of occlusive thrombi in stenotic arteries involves the rapid deposition of millions of circulating platelets under high shear flow. The process is mediated by the formation of molecular bonds of several distinct types between platelets; the bonds capture the moving platelets and stabilize the growing thrombi under flow. We investigated the mechanisms behind occlusive thrombosis in arteries with a two-phase continuum model. The model explicitly tracks the formation and rupture of the two types of interplatelet bonds, the rates of which are coupled with the local flow conditions. The motion of platelets in the thrombi results from competition between the viscoelastic forces generated by the interplatelet bonds and the fluid drag. Our simulation results indicate that stable occlusive thrombi form only under specific combinations for the ranges of model parameters such as rates of bond formation and rupture, platelet activation time, and number of bonds required for platelet attachment.

A model of fluid-structure and biochemical interactions for applications to subclinical leaflet thrombosis.

Subclinical leaflet thrombosis (SLT) is a potentially serious complication of aortic valve replacement with a bioprosthetic valve in which blood clots form on the replacement valve. SLT is associated with increased risk of transient ischemic attacks and strokes and can progress to clinical leaflet thrombosis. SLT following aortic valve replacement also may be related to subsequent structural valve deterioration, which can impair the durability of the valve replacement. Because of the difficulty in clinical imaging of SLT, models are needed to determine the mechanisms of SLT and could eventually predict which patients will develop SLT. To this end, we develop methods to simulate leaflet thrombosis that combine fluid-structure interaction and a simplified thrombosis model that allows for deposition along the moving leaflets. Additionally, this model can be adapted to model deposition or absorption along other moving boundaries. We present convergence results and quantify the model's ability to realize changes in valve opening and pressures. These new approaches are an important advancement in our tools for modeling thrombosis because they incorporate both adhesion to the surface of the moving leaflets and feedback to the fluid-structure interaction.

Towards understanding the effect of fibrinogen interactions on fibrin gel structure.

Fibrin gelation involves the enzymatic conversion of the plasma protein fibrinogen to fibrin monomers which then polymerize to form the gel that is a major structural component of a blood clot. Because fibrinogen provides the material from which fibrin is made, it is generally regarded as promoting the gelation process. However, fibrinogen can bind to a site on a fibrin oligomer, preventing another fibrin oligomer from binding there, thus slowing the polymerization process. "Soluble fibrin oligomers," which are mixtures of fibrin and fibrinogen, are found in the blood plasma and serve as biomarkers for various clotting disorders, so understanding the interplay between fibrin and fibrinogen during fibrin polymerization may have medical importance. We present a kinetic gelation model of fibrin polymerization which accounts for the dual and antagonistic roles of fibrinogen. It builds on our earlier model of fibrin polymerization that proposed a novel mechanism for branch formation, which is a necessary component of gelation. This previous model captured salient experimental observations regarding the determinants of the structure of the gel, but did not include fibrinogen binding. Here, we add to that model reactions between fibrinogen and fibrin, so oligomers are now mixtures of fibrin and fibrinogen, and characterizing their dynamics leads to equations of substantially greater complexity than previously. Using a moment generating function approach, we derive a closed system of moment equations and we track their dynamics until the finite time blow-up of specific second moments indicates that a gel has formed. In simulations begun with an initial mixture of fibrin and fibrinogen monomers, a sufficiently high relative concentration of fibrinogen prevents gelation; the critical concentration increases with the branch formation rate. In simulations begun with only fibrinogen monomers that are converted to fibrin at a specified rate, the rates of conversion, fibrinogen binding to oligomers, and branch formation together determine whether a gel forms, how long it takes to form, and the structural properties of the gel that results.

Inhibition of platelet-surface-bound proteins during coagulation under flow I: TFPI.

Blood coagulation is a self-repair process regulated by activated platelet surfaces, clotting factors, and inhibitors. Tissue factor pathway inhibitor (TFPI) is one such inhibitor, well known for its inhibitory action on the active enzyme complex comprising tissue factor (TF) and activated clotting factor VII. This complex forms when TF embedded in the blood vessel wall is exposed by injury and initiates coagulation. A different role for TFPI, independent of TF:VIIa, has recently been discovered whereby TFPI binds a partially cleaved form of clotting factor V (FV-h) and impedes thrombin generation on activated platelet surfaces. We hypothesized that this TF-independent inhibitory mechanism on platelet surfaces would be a more effective platform for TFPI than the TF-dependent one. We examined the effects of this mechanism on thrombin generation by including the relevant biochemical reactions into our previously validated mathematical model. Additionally, we included the ability of TFPI to bind directly to and inhibit platelet-bound FXa. The new model was sensitive to TFPI levels and, under some conditions, TFPI could completely shut down thrombin generation. This sensitivity was due entirely to the surface-mediated inhibitory reactions. The addition of the new TFPI reactions increased the threshold level of TF needed to elicit a strong thrombin response under flow, but the concentration of thrombin achieved, if there was a response, was unchanged. Interestingly, we found that direct binding of TFPI to platelet-bound FXa had a greater anticoagulant effect than did TFPI binding to FV-h alone, but that the greatest effects occurred if both reactions were at play. The model includes activated platelets' release of FV species, and we explored the impact of varying the FV/FV-h composition of the releasate. We found that reducing the zymogen FV fraction of this pool, and thus increasing the fraction that is FV-h, led to acceleration of thrombin generation.

Inhibition of platelet-surface-bound proteins during coagulation under flow II: Antithrombin and heparin.

Blood coagulation is a self-repair process regulated by activated platelet surfaces, clotting factors, and inhibitors. Antithrombin (AT) is one such inhibitor that impedes coagulation by targeting and inactivating several key coagulation enzymes. The effect of AT is greatly enhanced in the presence of heparin, a common anticoagulant drug. When heparin binds to AT, it either bridges with the target enzyme or induces allosteric changes in AT leading to more favorable binding with the target enzyme. AT inhibition of fluid-phase enzymes caused little suppression of thrombin generation in our previous mathematical models of blood coagulation under flow. This is because in that model, flow itself was a greater inhibitor of the fluid-phase enzymes than AT. From clinical observations, it is clear that AT and heparin should have strong inhibitory effects on thrombin generation, and thus we hypothesized that AT could be inhibiting enzymes bound to activated platelet surfaces that are not subject to being washed away by flow. We extended our mathematical model to include the relevant reactions of AT inhibition at the activated platelet surfaces as well as those for unfractionated heparin and a low molecular weight heparin. Our results show that AT alone is only an effective inhibitor at low tissue factor densities, but in the presence of heparin, it can greatly alter, and in some cases shut down, thrombin generation. Additionally, we studied each target enzyme separately and found that inactivation of no single enzyme could substantially suppress thrombin generation.

DEVELOPMENT OF FIBRIN BRANCH STRUCTURE BEFORE AND AFTER GELATION.

In [Fogelson and Keener, Phys. Rev. E, 81 (2010), 051922], we introduced a kinetic model of fibrin polymerization during blood clotting that captured salient experimental observations about how the gel branching structure depends on the conditions under which the polymerization occurs. Our analysis there used a moment-based approach that is valid only before the finite time blow-up that indicates formation of a gel. Here, we extend our analyses of the model to include both pre-gel and post-gel dynamics using the PDE-based framework we introduced in [Fogelson and Keener, SIAM J. Appl. Math., 75 (2015), pp. 1346-1368]. We also extend the model to include spatial heterogeneity and spatial transport processes. Studies of the behavior of the model reveal different spatial-temporal dynamics as the time scales of the key processes of branch formation, monomer introduction, and diffusion are varied.

A Hybrid Semi-Lagrangian Cut Cell Method for Advection-Diffusion Problems with Robin Boundary Conditions in Moving Domains.

We present a new discretization approach to advection-diffusion problems with Robin boundary conditions on complex, time-dependent domains. The method is based on second order cut cell finite volume methods introduced by Bochkov et al. [8] to discretize the Laplace operator and Robin boundary condition. To overcome the small cell problem, we use a splitting scheme along with a semi-Lagrangian method to treat advection. We demonstrate second order accuracy in the L 1, L 2, and L ∞ norms for both analytic test problems and numerical convergence studies. We also demonstrate the ability of the scheme to convert one chemical species to another across a moving boundary.

Modeling and Simulation of the Ion-Binding-Mediated Swelling Dynamics of Mucin-like Polyelectrolyte Gels.

Volume phase transitions in polyeletrolyte gels play important roles in many biophysical processes such as DNA packaging, nerve excitation, and cellular secretion. The swelling and deswelling of these charged polymer gels depend strongly on their ionic environment. In this paper, we present an extension to our previous two-fluid model for ion-binding-mediated gel swelling. The extended model eliminates the assumptions about the size similarity between the network and solvent particles, which makes it suitable for investigating of a large family of biologically relevant problems. The model treats the polyeletrolyte gel as a mixture of two materials, the network and the solvent. The dynamics of gel swelling is governed by the balance between the mechanical and chemical forces on each of these two materials. Simulations based on the model illustrate that the chemical forces are significantly influenced by the binding/unbinding reactions between the ions and the network, as well as the resulting distribution of charges within the gel. The dependence of the swelling rate on ionic bath concentrations is analyzed and this analysis highlights the importance of the electromigration of ions and the induced electric field in regulating gel swelling.

An Efficient High-Order Meshless Method for Advection-Diffusion Equations on Time-Varying Irregular Domains.

We present a high-order radial basis function finite difference (RBF-FD) framework for the solution of advection-diffusion equations on time-varying domains. Our framework is based on a generalization of the recently developed Overlapped RBF-FD method that utilizes a novel automatic procedure for computing RBF-FD weights on stencils in variable-sized regions around stencil centers. This procedure eliminates the overlap parameter δ, thereby enabling tuning-free assembly of RBF-FD differentiation matrices on moving domains. In addition, our framework utilizes a simple and efficient procedure for updating differentiation matrices on moving domains tiled by node sets of time-varying cardinality. Finally, advection-diffusion in time-varying domains is handled through a combination of rapid node set modification, a new high-order semi-Lagrangian method that utilizes the new tuning-free overlapped RBF-FD method, and a high-order time-integration method. The resulting framework has no tuning parameters and has O(N logN) time complexity. We demonstrate high-orders of convergence for advection-diffusion equations on time-varying 2D and 3D domains for both small and large Peclet numbers. We also present timings that verify our complexity estimates. Finally, we utilize our method to solve a coupled 3D problem motivated by models of platelet aggregation and coagulation, once again demonstrating high-order convergence rates on a moving domain.

Pump efficacy in a two-dimensional, fluid-structure interaction model of a chain of contracting lymphangions.

The transport of lymph through the lymphatic vasculature is the mechanism for returning excess interstitial fluid to the circulatory system, and it is essential for fluid homeostasis. Collecting lymphatic vessels comprise a significant portion of the lymphatic vasculature and are divided by valves into contractile segments known as lymphangions. Despite its importance, lymphatic transport in collecting vessels is not well understood. We present a computational model to study lymph flow through chains of valved, contracting lymphangions. We used the Navier-Stokes equations to model the fluid flow and the immersed boundary method to handle the two-way, fluid-structure interaction in 2D, non-axisymmetric simulations. We used our model to evaluate the effects of chain length, contraction style, and adverse axial pressure difference (AAPD) on cycle-mean flow rates (CMFRs). In the model, longer lymphangion chains generally yield larger CMFRs, and they fail to generate positive CMFRs at higher AAPDs than shorter chains. Simultaneously contracting pumps generate the largest CMFRs at nearly every AAPD and for every chain length. Due to the contraction timing and valve dynamics, non-simultaneous pumps generate lower CMFRs than the simultaneous pumps; the discrepancy diminishes as the AAPD increases. Valve dynamics vary with the contraction style and exhibit hysteretic opening and closing behaviors. Our model provides insight into how contraction propagation affects flow rates and transport through a lymphangion chain.

The Art and Science of Building a Computational Model to Understand Hemostasis.

Computational models of various facets of hemostasis and thrombosis have increased substantially in the last decade. These models have the potential to make predictions that can uncover new mechanisms within the complex dynamics of thrombus formation. However, these predictions are only as good as the data and assumptions they are built upon, and therefore model building requires intimate coupling with experiments. The objective of this article is to guide the reader through how a computational model is built and how it can inform and be refined by experiments. This is accomplished by answering six questions facing the model builder: (1) Why make a model? (2) What kind of model should be built? (3) How is the model built? (4) Is the model a "good" model? (5) Do we believe the model? (6) Is the model useful? These questions are answered in the context of a model of thrombus formation that has been successfully applied to understanding the interplay between blood flow, platelet deposition, and coagulation and in identifying potential modifiers of thrombin generation in hemophilia A.

Computationally Driven Discovery in Coagulation.

Bleeding frequency and severity within clinical categories of hemophilia A are highly variable and the origin of this variation is unknown. Solving this mystery in coagulation requires the generation and analysis of large data sets comprised of experimental outputs or patient samples, both of which are subject to limited availability. In this review, we describe how a computationally driven approach bypasses such limitations by generating large synthetic patient data sets. These data sets were created with a mechanistic mathematical model, by varying the model inputs, clotting factor, and inhibitor concentrations, within normal physiological ranges. Specific mathematical metrics were chosen from the model output, used as a surrogate measure for bleeding severity, and statistically analyzed for further exploration and hypothesis generation. We highlight results from our recent study that employed this computationally driven approach to identify FV (factor V) as a key modifier of thrombin generation in mild to moderate hemophilia A, which was confirmed with complementary experimental assays. The mathematical model was used further to propose a potential mechanism for these observations whereby thrombin generation is rescued in FVIII-deficient plasma due to reduced substrate competition between FV and FVIII for FXa (activated factor X).

Clot Permeability, Agonist Transport, and Platelet Binding Kinetics in Arterial Thrombosis.

The formation of wall-adherent platelet aggregates is a critical process in arterial thrombosis. A growing aggregate experiences frictional drag forces exerted on it by fluid moving over or through the aggregate. The magnitude of these forces is strongly influenced by the permeability of the developing aggregate; the permeability depends on the aggregate's porosity. Aggregation is mediated by formation of ensembles of molecular bonds; each bond involves a plasma protein bridging the gap between specific receptors on the surfaces of two different platelets. The ability of the bonds existing at any time to sustain the drag forces on the aggregate determines whether it remains intact or sheds individual platelets or larger fragments (emboli). We investigate platelet aggregation in coronary-sized arteries using both computational simulations and in vitro experiments. The computational model tracks the formation and breaking of bonds between platelets and treats the thrombus as an evolving porous, viscoelastic material, which moves differently from the background fluid. This relative motion generates drag forces which the fluid and thrombus exert on one another. These forces are computed from a permeability-porosity relation parameterized from experimental measurements. Basing this relation on measurements from occlusive thrombi formed in our flow chamber experiments, along with other physiological parameter values, the model produced stable dense thrombi on a similar timescale to the experiments. When we parameterized the permeability-porosity relation using lower permeabilities reported by others, bond formation was insufficient to balance drag forces on an early thrombus and keep it intact. Under high shear flow, soluble agonist released by platelets was limited to the thrombus and a boundary layer downstream, thus restricting thrombus growth into the vessel lumen. Adding to the model binding and activation of unactivated platelets through von Willebrand-factor-mediated processes allowed greater growth and made agonist-induced activation more effective.

Kinetic model of two-monomer polymerization.

We propose a kinetic gelation model of polymer growth with two monomeric types that have distinct functionalities (reaction sites), and can polymerize using different reaction types. The heterotypic aggregation of two monomer types is modeled using a moment generating function approach by tracking the temporal evolution of a closed system of moment equations up until gelation. We investigate several scenarios of polymerization with two distinct monomers that differ in the types of reactions that can occur. We determine numerical and analytical conditions for finite time blow-up (the emergence of an oligomer of infinite size) that depend on initial conditions, reaction rates, and number of reaction sites per monomer.

A mathematical model of coagulation under flow identifies factor V as a modifier of thrombin generation in hemophilia A.

BACKGROUND: The variability in bleeding patterns among individuals with hemophilia A, who have similar factor VIII (FVIII) levels, is significant and the origins are unknown. OBJECTIVE: To use a previously validated mathematical model of flow-mediated coagulation as a screening tool to identify parameters that are most likely to enhance thrombin generation in the context of FVIII deficiency. METHODS: We performed a global sensitivity analysis (GSA) on our mathematical model to identify potential modifiers of thrombin generation. Candidates from the GSA were confirmed by calibrated automated thrombography (CAT) and flow assays on collagen-tissue factor (TF) surfaces at a shear rate of 100 per second. RESULTS: Simulations identified low-normal factor V (FV) (50%) as the strongest modifier, with additional thrombin enhancement when combined with high-normal prothrombin (150%). Low-normal FV levels or partial FV inhibition (60% activity) augmented thrombin generation in FVIII-inhibited or FVIII-deficient plasma in CAT. Partial FV inhibition (60%) boosted fibrin deposition in flow assays performed with whole blood from individuals with mild and moderate FVIII deficiencies. These effects were amplified by high-normal prothrombin levels in both experimental models. CONCLUSIONS: These results show that low-normal FV levels can enhance thrombin generation in hemophilia A. Further explorations with the mathematical model suggest a potential mechanism: lowering FV reduces competition between FV and FVIII for factor Xa (FXa) on activated platelet surfaces (APS), which enhances FVIII activation and rescues thrombin generation in FVIII-deficient blood.

A MATHEMATICAL MODEL OF PLATELET AGGREGATION IN AN EXTRAVASCULAR INJURY UNDER FLOW.

We present the first mathematical model of flow-mediated primary hemostasis in an extravascular injury which can track the process from initial deposition to occlusion. The model consists of a system of ordinary differential equations (ODEs) that describe platelet aggregation (adhesion and cohesion), soluble-agonist-dependent platelet activation, and the flow of blood through the injury. The formation of platelet aggregates increases resistance to flow through the injury, which is modeled using the Stokes-Brinkman equations. Data from analogous experimental (microfluidic flow) and partial differential equation models informed parameter values used in the ODE model description of platelet adhesion, cohesion, and activation. This model predicts injury occlusion under a range of flow and platelet activation conditions. Simulations testing the effects of shear and activation rates resulted in delayed occlusion and aggregate heterogeneity. These results validate our hypothesis that flow-mediated dilution of activating chemical adenosine diphosphate hinders aggregate development. This novel modeling framework can be extended to include more mechanisms of platelet activation as well as the addition of the biochemical reactions of coagulation, resulting in a computationally efficient high throughput screening tool of primary and secondary hemostasis.

A local and global sensitivity analysis of a mathematical model of coagulation and platelet deposition under flow.

The hemostatic response involves blood coagulation and platelet aggregation to stop blood loss from an injured blood vessel. The complexity of these processes make it difficult to intuit the overall hemostatic response without quantitative methods. Mathematical models aim to address this challenge but are often accompanied by numerous parameters choices and thus need to be analyzed for sensitivity to such choices. Here we use local and global sensitivity analyses to study a model of coagulation and platelet deposition under flow. To relate with clinical assays, we measured the sensitivity of three specific thrombin metrics: lag time, maximum relative rate of generation, and final concentration after 20 minutes. In addition, we varied parameters of three different classes: plasma protein levels, kinetic rate constants, and platelet characteristics. In terms of an overall ranking of the model's sensitivities, we found that the local and global methods provided similar information. Our local analysis, in agreement with previous findings, shows that varying parameters within 50-150% of baseline values, in a one-at-a-time (OAT) fashion, always leads to significant thrombin generation in 20 minutes. Our global analysis gave a different and novel result highlighting groups of parameters, still varying within the normal 50-150%, that produced little or no thrombin in 20 minutes. Variations in either plasma levels or platelet characteristics, using either OAT or simultaneous variations, always led to strong thrombin production and overall, relatively low output variance. Simultaneous variation in kinetics rate constants or in a subset of all three parameter classes led to the highest overall output variance, incorporating instances with little to no thrombin production. The global analysis revealed multiple parameter interactions in the lag time and final concentration leading to relatively high variance; high variance was also observed in the thrombin generation rate, but parameters attributed to that variance acted independently and additively.

A Two-phase mixture model of platelet aggregation.

We present a two-phase model of platelet aggregation in coronary-artery-sized blood vessels. The model tracks the number densities of three platelet populations as well as the concentration of a platelet activating chemical. Through the formation of elastic bonds, activated platelets can cohere with one another to form a platelet thrombus. Bound platelets in a thrombus move in a velocity field different from that of the bulk fluid. Stresses produced by the elastic bonds act on the bound platelet material. Movement of the bound platelet material and that of the background fluid are coupled through an interphase drag and an incompressibility constraint. The relative motion between bound platelets and the background fluid permits intraclot transport of individual platelets and activating chemical, allows the bound platelet density to reach levels much higher than the platelet density in the bulk blood, and allows thrombus formation to occur on a physiological timescale, all of which were precluded by our earlier single phase model. Computational results from the two-phase model indicate that through complicated fluid-structure interactions, the platelet thrombus can develop significant spatial inhomogeneities and that the amount of intraclot flow may greatly affect the growth, density, and stability of a thrombus.

Electrodiffusion-Mediated Swelling of a Two-Phase Gel Model of Gastric Mucus.

Gastric mucus gel is known to exhibit dramatic and unique swelling behaviors in response to the ionic composition of the hydrating solution. This swelling behavior is important in the maintenance of the mucus layer lining the stomach wall, as the layer is constantly digested by enzymes in the lumen, and must be replenished by new mucus that swells as it is secreted from the gastric wall. One hypothesis suggests that the condensed state of mucus at secretion is maintained by transient bonds with calcium that form crosslinks. These crosslinks are lost as monovalent cations from the environment displace divalent crosslinkers, leading to a dramatic change in the energy of the gel and inducing the swelling behavior. Previous modeling work has characterized the equilibrium behavior of polyelectrolyte gels that respond to calcium crosslinking. Here, we present an investigation of the dynamic swelling behavior of a polyelectrolytic gel model of mucus. In particular, we quantified the rate at which a globule of initially crosslinked gel swells when exposed to an ionic bath. The dependence of this swelling rate on several parameters was characterized. We observed that swelling rate has a non-monotone dependence on the molarity of the bath solution, with moderate concentrations of available sodium inducing the fastest swelling.

Hyperviscosity-Based Stabilization for Radial Basis Function-Finite Difference (RBF-FD) Discretizations of Advection-Diffusion Equations.

We present a novel hyperviscosity formulation for stabilizing RBF-FD discretizations of the advectiondiffusion equation. The amount of hyperviscosity is determined quasi-analytically for commonly-used explicit, implicit, and implicit-explicit (IMEX) time integrators by using a simple 1D semi-discrete Von Neumann analysis. The analysis is applied to an analytical model of spurious growth in RBF-FD solutions that uses auxiliary differential operators mimicking the undesirable properties of RBF-FD differentiation matrices. The resulting hyperviscosity formulation is a generalization of existing ones in the literature, but is free of any tuning parameters and can be computed efficiently. To further improve robustness, we introduce a simple new scaling law for polynomial-augmented RBF-FD that relates the degree of polyharmonic spline (PHS) RBFs to the degree of the appended polynomial. When used in a novel ghost node formulation in conjunction with the recently-developed overlapped RBF-FD method, the resulting method is robust and free of stagnation errors. We validate the high-order convergence rates of our method on 2D and 3D test cases over a wide range of Peclet numbers (1-1000). We then use our method to solve a 3D coupled problem motivated by models of platelet aggregation and coagulation, again demonstrating high-order convergence rates.