Multiscale Modeling of Blood Flow-Mediated Platelet Thrombosis
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The blood coagulation cascade that leads to thrombus formation may be initiated by flow-induced platelet activation, which prompts clot formation in prosthetic cardiovascular devices and in arterial disease processes. Upon activation, platelets undergo complex morphological changes of filopodia formation that play a major role in aggregation and attachment to surfaces. Numerical simulations based on continuum approaches fail to capture such molecular-scale mechano-transduction processes. Utilizing molecular dynamics (MD) to model these complex processes across the scales is computationally prohibitive. We describe multiscale numerical methodologies that integrate four key components of blood clotting, namely, blood rheology, cell mechanics, coagulation kinetics and transport of species, and platelet adhesive dynamics across a wide range of spatiotemporal scales. Whereas mechanics of binding/unbinding for single-molecule receptor-ligand complexes can be simulated by molecular dynamics (MD), the mechanical structure of platelets in blood flow and their interaction with flow-induced stresses that may lead to their activation can be efficiently described by a model at coarser scales, using numerical approaches such as coarse-grained molecular dynamics (CGMD). Additionally, CGMD provides an excellent platform to inform other coarser-scale models in a bottom-up approach in the multiscale hierarchy. The microenvironment of most biological systems such as coagulation normally involves a large number of cells, e.g., blood cells suspended in plasma, limiting the utility of CGMD at the larger transport scales of blood flow. However, dissipative particle dynamics (DPD), along with its sub-models such as energy conserving and transport DPD, provides a very flexible platform for scaling up these mesoscopic systems. At the macroscopic top scales of the vasculature and cardiovascular devices, simulating blood and tissues using continuum-based methods becomes viable and efficient. However, the challenge of interfacing these larger transport scales with the orders of magnitude smaller spatiotemporal scales that characterize blood coagulation, and given the issue of the slow-dynamic timescales of biological processes, makes long-term simulations of such systems computationally prohibitive. In this chapter, we describe various numerical remedies based on these methodologies that facilitate overcoming this multiscale simulation challenge.
KeywordsMultiscale Modeling Coarse-grained Molecular Dynamics (CGMD) Dissipative Particle Dynamics (DPD) Mechano-transduction Process Flow-induced Stresses
Hemostasis (thrombus formation) is the normal physiological response that prevents significant blood loss after vascular injury. The resulting clots can form under different flow conditions in the veins as well as the arteries. The excessive and undesirable formation of clots (i.e., thrombosis) in our circulatory system may lead to significant morbidity and mortality. Some of these pathologies are deep vein thrombosis and pulmonary embolism and atherothrombosis (thrombosis triggered by plaque rupture) in coronary arteries, to name a few. The process of clot formation and growth at a site on a blood vessel wall involves a number of simultaneous processes including multiple chemical reactions in the coagulation cascade, species transport, and platelet adhesion, all of which are strongly influenced by the hydrodynamic forces.
While platelet activation may be induced by biochemical agonists, shear stresses arising from pathological flow patterns enhance the propensity of platelets to activate and initiate coagulation cascade, causing thrombosis. Flow stresses acting on the platelets can be represented down to the order of microns by continuum models. However, processes such as the morphological changes in platelets upon activation happen at the molecular scales. Continuum approaches fail to capture such processes. Mesoscopic particle-based methods were developed to model blood cells and thrombosis in flowing blood, using dissipative particle dynamics (DPD) for modeling the blood, blood flows in microvessel, interactions of cells, and thrombosis growth. However, complex processes such as platelet activation require drastically different modeling approaches in order to achieve highly resolved microstructural mechanics and the mechano-transduction process of hemodynamic stresses that may induce platelet activation. Such approaches are needed for more accurately describing the platelets intracellular structure and the mechano-transduction processes of platelet activation by hemodynamic forces.
Incorporating all these processes that occur at different spatiotemporal scales, however, remains a rather challenging task. For example, platelet-wall and platelet-platelet interactions through receptor-ligand bindings occur at a subcellular nanoscale, whereas the blood flow dynamics in the vessel around the developing thrombus is described as a macroscopic process from several micrometers to millimeters. Here, we describe a hierarchical multiscale numerical approach that may seamlessly unify and integrate different subprocesses within the clotting process.
1.1 Significance and Rationale for a Multiscale Model
Cardiovascular diseases account for over 31% of deaths globally and in the USA annually. Whether due to acute thrombosis associated with myocardial infarction or progressive intermittent atherothrombotic events, significant ventricular dysfunction may result, leading to heart failure. Presently over 6.5 million patients suffer from heart failure in USA, and their number is expected to grow by nearly 50% by 2030. Of those, a significant proportion will become candidates for mechanical circulatory support and prosthetic cardiovascular devices, also burdened with thromboembolic risk and complications, mandating antithrombotic medications for their recipients. Despite antithrombotic therapy, ventricular assist device patients have approximately a 12% stroke rate, of which 18% prove fatal. The alarming thrombosis rates, coupled with complex antithrombotic and anticoagulant management, necessitate a solution to save numerous lives and drastically reduce the ensuing healthcare costs.
Thrombosis in vascular disease is potentiated by the interaction of blood constituents with an injured vascular wall and the nonphysiologic flow patterns characterizing cardiovascular pathologies. Both initiate and enhance the hemostatic response through chronic platelet activation. Similarly, the thrombus and thromboemboli generated in devices are composed primarily of platelets. The elevated flow stresses they induce chronically activate platelets despite aggressive antithrombotic therapy. However, limitations in characterizing the mechanical stimuli, and the complex biochemical and morphological changes platelets undergo in response, still hamper our ability to successfully model such intricate phenomena. The coupling of the disparate spatiotemporal scales between molecular-level events and the macroscopic transport represents a major modeling and computational challenge, which requires a multidisciplinary integrated multiscale numerical approach. Continuum approaches are limited in their ability to cover the smaller molecular mechanisms such as filopodia formation during platelet activation. Utilizing molecular dynamics (MD) to cover the multiscales involved is computationally prohibitive. Innovative multiscale approaches are essential for elucidating this vexing problem (Bluestein et al. 2014; Zhang et al. 2014).
1.2 Flow-Induced Mechanisms of Platelet Activation, Aggregation, and Thrombosis
Low-intermediate shear in veins and larger arteries (<103 s−1). Aggregation is predominantly mediated by αIIbβ3 on free-flowing platelets which engage fibrinogen (Fg) adsorbed on platelet monolayers that compose the initial thrombi.
High shear in arterial microcirculation or moderate arterial stenosis (103 − 104 s−1). Aggregation is more vWF-dependent and involves the activity of both GPIba and αIIbβ3.
Pathological or “hyper” shear in atherothrombosis and cardiovascular devices (>104 s−1). Aggregation is mediated exclusively by vWF-GPIb bonds, not requiring platelet activation or adhesion via αIIbβ3.
1.3 Aggregation, Adhesion, and Wall Interaction
Shear-induced flow mechanisms are enhanced by eddies, flow vortices, and flow separation (Nesbitt et al. 2009), with prothrombotic agonists trapped in regions of recirculating flow. Shear-induced aggregation has two characteristic phases: (i) reversible phase between two discoid platelets and the presence of unstable platelet aggregates and (ii) stable phase that involves platelet shape change and granular release (Maxwell et al. 2007; Nesbitt et al. 2009). Unstable aggregates are formed with vWF alone, while stable aggregates require both vWF and fibrinogen, allowing for a shorter formation time and increased recruitment of platelets to aggregates (Maxwell et al. 2007). Initial aggregates typically involve clustering of platelets around a single activated platelet via membrane tethers, not pseudopodia, whereas stable aggregates require platelet sphering and filopodial projections. Increasing the shear rate from 1,800 to 5,000 s−1 increases the number of discoid platelet aggregates converting to stable aggregates, and αIIbβ3 is required for membrane tether formation between platelets in reversible aggregates (Maxwell et al. 2007). In stenotic regions, initial platelet recruitment occurs at the stenosis apex, and aggregates form at the downstream expansion zone where the flow decelerates. Larger decelerations result in larger aggregates embedded in the growing thrombus.
1.4 Continuum Approaches for Blood Flow at the Microscale
Several continuum approaches try to couple the timescales between molecular and macroscopic transport. In those the cellular level is represented by field equations and moving boundaries to allow the cell to change its shape with interfaces that have different timescales and properties (N’Dri et al. 2003; Shyy et al. 2001). Models of, e.g., adhesion of rolling leukocytes to endothelium are based on either equilibrium concepts (Bell 1978; Berk and Evans 1991; Evans 1985a,b) or kinetics approach (Dembo et al. 1988; Hammer and Lauffenburger 1987; Dong et al. 1999). Bond association and dissociation occur according to the forward and reverse reaction rates (N’Dri et al. 2003; Shyy et al. 2001). Initial models (Skalak et al. 1981a, 1989, 1981b; Weiss 1990) neglected cell rheological properties and were limited in their predictive capability (Bell 1978; Dembo et al. 1988). Flow-induced stresses acting on a cell may weaken the adhesion bonds or extract receptor molecules from the cell surface (Alon et al. 1997, 1998) with progress toward understanding the receptor-mediated cell adhesion process (Lauffenburger et al. 1987; Linderman and Lauffenburger 1986, 1988). Detailed review on such simulations using probabilistic and Monte Carlo approaches was published (Zhu 2000). Flow-mediated transport and initiation and inhibition of coagulation were modeled extensively by the Fogelson group (Fogelson and Tania 2005).
1.5 Coarse-Grained Particle Methods at the Mesoscale
CFD simulations are limited to continuum systems (Dwinzel et al. 2002; Groot and Warren 1997), representing a 10 nm to 100 μm gap between microscopic biochemistry and macroscopic scales. DPD models possess important properties of mesoscopic systems: DPD can easily model heterogeneous fluids, allowing simulations of processes which are very difficult to model by continuum approaches. Initial work focused on multiphase nature of these flows (Espanol 1998; Boek et al. 1996) to address limitations of continuum approaches that represent them consistently (Albert et al. 1998; Kechagia et al. 2002; Davis 2002). Blood flow was modeled with DPD (Dzwinel et al. 2003) with plasma represented by dissipative fluid particles, and red blood cells (RBCs) and capillary walls by coarse-grained (CG) particles ensemble (Boryczko et al. 2003, 2004), demonstrating aggregation and deformation. A triple-decker MD-DPD model interfaced atomistic-mesoscopic-continuum flow regimes (Fedosov and Karniadakis 2009). A flow-cell interaction model (Sweet et al. 2011) used subcellular elements to simulate cell motion, with the cell coupled to the plasma flow by the Langevin equation (Skeel and Izaguirre 2002). DPD was initially used in simple geometries (Fedosov and Karniadakis 2009; Soddemann et al. 2003; Espanol and Warren 1995) for modeling viscous flows with no-slip boundary conditions (Koplik and Banavar 1995a,b; Revenga et al. 1999; Pivkin and Karniadakis 2005), expanding to higher Reynolds numbers (Qian and Wang 2005), and more recently in a stenosis model (Gao et al. 2017).
2 Hierarchical Multiscale Modeling of Platelet Thrombosis
Numerical modeling of thrombus formation and growth is a challenging problem due to multiscale and multiphysics nature of clotting process, which involves fluid mechanics, cell mechanics, and biochemistry. Diverse studies have addressed this problem on different scales such as cellular, meso, and continuum levels, whereas attempts have been made to bridge these different scales to model the process at the initial phase of platelet activation and aggregation (cf. Xu et al. 2010, Zhang et al. 2014, and Grinberg et al. 2013). These studies may be broadly put in three distinct modeling strategies: cellular/subcellular modeling of adhesive dynamics, platelet transport, and aggregation in whole blood; continuum-based modeling of blood flow treating platelets as Lagrangian particles; and continuum-based modeling of thrombus formation and growth using empirical correlations for platelet deposition rates.
2.1 Molecular Dynamics Modeling of Receptor-Ligand Interactions
Atomic force microscope (AFM) experiments have been used extensively to probe the force required to rupture single-molecule complexes and have provided additional insights into their binding properties. By performing force spectroscopic measurements of binding/unbinding force distributions at different relaxation/pulling rates, these AFM experiments have been able to quantify the kinetics of ligand-receptor interactions.
To reveal the microscopic processes underlying the AFM observations, numerical simulations of binding/unbinding events for different complexes have been performed using steered molecular dynamics (SMD) technique. SMD allows us to explore these processes on timescales accessible to MD simulations. The basic idea behind any SMD simulation is to apply an external force to one or more atoms. In addition, you can keep another group of atoms fixed and study the behavior of the complex under various conditions. The only major difference between the AFM experiment and simulations concerns the value for the pulling velocity. Whereas the AFM experiment is carried out on a millisecond timescale, MD simulations are limited to nanoseconds; therefore, one has to consider thermal fluctuations and dissipation. Because of these nonequilibrium phenomena, the rupture force should vary systematically with rupture speed, and thus the computed rupture forces should be able to be extrapolated to the experimental timescale (Grubmüller et al. 1996).
2.2 An Integrated DPD-CGMD Modeling Approach
2.2.1 Coarse-Grained Molecular Dynamics Model of Platelet Cellular Structure
The cytoplasm rheology is modeled using a Morse potential (Morse 1929). The cytoskeleton consists of two types of actin-based filaments: (a) a rigid filamentous core and (b) an assembly of radially spanning elastic actin filaments that mediates the contractility. A carbon-70 structure is used to generate the oval-shaped core. Each actin filament is individually extensible and is tethered to the core. An α-helical structure mimics the spring-loaded molecular mechanism, with its spiral conformation continuously stretchable under external force.
2.2.2 Modeling Blood Plasma Using DPD Fluid Model
2.2.3 DPD-CGMD Spatial Interfacing
Determination of the parameters for the spatial interface is conducted through numerical experiments for maintaining the Jefferys orbits of spheroids in a shear flow. The Jefferys orbits solution (Jeffery 1922) describes the rotation of isolated ellipsoidal particles immersed in viscous shear flow and is widely used as a benchmark analytical solution. Zhang et al. (2014) presented the comparison of this multiscale fluid-platelet simulation results with Jeffery’s orbit solution in Couette flows.
2.2.4 Modeling the Highly Resolved Mechano-Transduction Process of Hemodynamic Stresses in Platelets
2.2.5 Modeling the Activation of Platelets and Formation of Filopodia
Flow-mediated platelet shape change can be induced by exposing the platelets to a range of physiologic and pathologic shear stresses encountered during blood flow. Platelets isolated by gel filtration are diluted in Hepes-buffered modified Tyrodes solution with 0.1% bovine serum albumin, recalcified with a physiological concentration of CaCl2, and exposed to shear stresses ranging from the physiologic 1 dyne∕cm2 to the pathologic 70 dyne∕cm2 for up to 4 mins in the hemodynamic shearing device (HSD), a programmable cone-plate-Couette viscometer that exposes cell suspensions to uniform dynamic shear stresses (Sheriff et al. 2013). During exposure, samples are withdrawn using a LabView-controlled syringe-capillary viscometer and immediately fixed in 1% paraformaldehyde to block additional shape change (Pothapragada et al. 2015). Fixed platelets are then placed on glass slides precoated with poly-L-lysine, dehydrated through a graded ethanol series, and sputter-coated with gold particles prior to imaging. Images are obtained at 30,000x magnification in a SEM and analyzed using a MATLAB image processing toolbox (Pothapragada et al. 2015). Platelet boundary parameters, such as the major and minor axis lengths, and circularity are measured by fitting an ellipse to the platelet perimeter, while filopod thickness is obtained by manually defining the intersection with the elliptical platelet body. Filopod lengths are then obtained by morphological erosion of the central filopod axis. These parameters are then recorded in a database and grouped by shear condition (i.e., shear stress and exposure time). The analyzed images are compared with snapshots of the simulation results in Fig. 8. The lengths of filopodia generated due to shear stresses in this study range from 0.24 to 2.74 μm and thicknesses range from 0.06 to 0.73 μm, and the simulations are able to achieve results within this range (Pothapragada et al. 2015).
2.2.6 Measuring Platelet Stiffness and Deformation with Dielectrophoresis-Mediated Electro-Deformation
2.3 Seamless Multiscale Modeling of Thrombosis Using Dissipative Particle Dynamics
Dissipative particle dynamics, a mesoscopic particle-based hydrodynamics approach, has been effectively used to model plasma and suspending cells in blood in many applications. The advantage is the seamless integration of hydrodynamics and cell mechanics in a single framework. Further, a new submodel of DPD that is able to address chemical transport has been formulated (Li et al. 2015). The multiscale model of thrombosis includes four main processes: the hemodynamics, coagulation kinetics and transport of species, blood cell mechanics, and platelet adhesive dynamics. DPD can provide the correct hydrodynamic behavior of fluids at the mesoscale, and it has been successfully applied to study blood, where a coarse-grained representation of the cell membrane is used for red blood cells and platelets.
2.4 Continuum and Particle/Continuum Modeling of Thrombosis
3 Concurrent Coupling Using a Domain Decomposition Approach
The bridging of scales is achieved through designing appropriate interface conditions between the heterogeneous solvers. The interface conditions are derived to respect the physical conservation constraints and the requirements of each solver. Specifically, mass conservation is of primary importance, while momentum and energy conservation are desirable but present several challenges, with the latter being nearly impossible in practice. Interface conditions are concurrently enforced by constructing appropriate projection, interpolation, denoising, and statistical moment-matching techniques that aim to transfer information across the different domains in a compatible way with the requirements and data format of each solver.
Coupling of atomistic and continuum solvers requires the calculation and communication of averaged properties, such as fluid velocity and density, across heterogeneous solver interfaces. For example, atomistic solvers require a local velocity flux to be imposed at each voxel of the atomistic domain. This is achieved through constructing appropriate interpolation and projection operators that are capable of mapping the continuum velocity field onto the atomistic domain. Specifically, to enforce mass conservation, the continuum solver computes the fluxes through the surface interfaces with the atomistic domains, and particles are inserted in the atomistic domain in such a way that these fluxes are preserved.
Accurate computing of averaged fields through processing of non-stationary atomistic data presents several challenges including geometrical complexity of the atomistic domains, thermal fluctuations, and flow unsteadiness. Therefore, effective averaging and filtering techniques are required before the data are transmitted to the continuum domains. In particular, to filter out the stochastic component (thermal fluctuations) at all interfaces with the continuum domain, a window proper orthogonal decomposition (WPOD) has been used and shown to be very effective (Grinberg et al. 2014).
4 Multiple Time-Stepping (MTS) Algorithm for Efficient Multiscale Modeling
The disparate spatial scales between the DPD flow and the CGMD platelet model are handled by a hybrid force field. However, the disparity in temporal scales between the two models represents a huge challenge that may become prohibitive in the practice of multiscale modeling. A multiple time-stepping (MTS) algorithm is the necessary solution for this kind of multiscale models. Conventional MTS algorithms manage to differentiate multi-stepping for up to one order of magnitude of scale (Han et al. 2007) and are being applied to classical MD simulations. In order to handle three to four orders of magnitude disparity in the temporal scales between DPD and CGMD, a novel improved MTS scheme hybridizing DPD and CGMD is developed through utilizing four different time-stepping sizes. In this MTS scheme, the DPD-based fluid system advances at the largest time step, the fluid-platelet interface at a middle time step size, and the non-bonded and bonded potentials for the CGMD-based platelet system at two smallest time step sizes (Zhang et al. 2015).
MTS algorithm is a trade-off between speed and accuracy for multiscale modeling. Energy conservation and maintenance of adequate precisions of other measures must be verified while accelerating computations. We investigate the microscopic shape changes of platelets in response to macroscopic flow-induced stresses. Thus, in measuring numerical solutions, we focus on the accuracy of characterizing the hybrid system and the flowing platelets, as well as the accuracy of calculating the dynamic flow-induced stresses on the surface membrane. To compare, the standard time stepping (STS) that operates a single time step size is examined against MTS. The numerical results demonstrated that the microscopic measures for single platelets are more sensitive to the MTS parameters than the macroscopic measures for the hybrid system. The results reaffirm that a separation of temporal scales in MTS considerably improves the efficiency of utilizing parallel computing resources, as compared to conventional single-scale methods in which considerable time is wasted conducting massive unnecessary computations. For example, completing 1-ms multiscale simulation of a 10-million particle system is reduced from 2.6 years to 3.5 days only (Zhang et al. 2015, 2016).
5 Comments on Long-Term Modeling of Thrombus Formation
In contrast to the concurrent coupling, serial coupling schemes determine an effective macroscale model from the microscale model in a preprocessing step and use the resulted macroscale model subsequently. Such serial coupling methods are largely limited to parameter passing yet are very useful in addressing long-term biological processes provided there is a timescale separation between its relevant subprocesses. For example, in Fig. 12, a possible strategy to tackle the long-term simulation of clot formation and remodeling upon arterial wall injury that normally occurs over days is proposed. Here, the subprocesses are identified as (i) the acute formation of thrombus and platelet aggregation over the first tens of seconds, (ii) the midterm formation of fibrin network at the site of thrombus and stabilization of the clot, and (iii) the long-term remodeling of the clot, which occurs over days. As noted in the figure, due to the separation of timescales between phases (i) and (ii), a serial coupling between the fine-scale particle/continuum (based on FCM) and the coarse-scale multiphasic continuum models is possible. The challenge is the long-term simulation of particles actively coupled with flow; the complexities in geometry and flow conditions and the large size of injuries may require hundreds of thousands of FCM particles to represent platelets, which imposes a restrictively high computational cost for such simulations in large domains. To overcome this difficulty, it is possible to initially distribute particles in the regions of interest that contain thrombogenic surfaces. The timescales of phases (ii) and (iii) are not separated, which implicates a concurrent coupling scheme between these two subprocesses.
This work was supported by NIH grants U01HL116323 (Yazdani, A.), NHLBI R21HL096930-01, NIBIB Quantum U01EB012487, and NHLBI U01HL131052 (Bluestein, D.) and XSEDE grants DMS140019, DMS150011 (Zhang, P.), and DMS140007 (Yazdani, A.).
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