Shear-induced Platelet Activation Research Articles

The impact of the Impeller's hub design on the performance and blood damage in a microaxial mechanical circulatory support device – A numerical study

This study uses CFD methods to investigate the effects of the impeller's geometry on the hemodynamic characteristics, pump performance, and blood damage parameters, in a percutaneous microaxial Mechanical Circulatory Support (MCS) device. The numerical simulations employ the steady state Reynolds-Averaged Navier-Stokes approximation using the SST k−ω turbulent model. Three different impeller models are examined with different hub conversion angles (α = 0○, 3○ and 5○). The analysis includes 23 cases for different pressure heads (Δp = 60–80 mmHg) and angular velocities (ω = 30–52 kRPM). The obtained flow rate is compared between the cases to assess the effect of the impeller's design and working conditions on the pump performance. The comparative risk of shear-induced platelet activation is estimated using the statistical median of the stress-accumulation values calculated along streamlines. The risk of hemolysis is estimated using the average exposure time to shear stress above a threshold (τ > 425 Pa). The results reveal that the shape of the impeller's hub has a great impact on the flow patterns, performance, and risk of blood damage, as well as the angular velocity. The highest flow rate (Q = 3.7 L/min) and efficiency (η = 11.3 %) were achieved using a straight hub (α = 0○). Similarly, for the same condition of flow and pressure, the straight hub impeller has the lowest blood damage risk parameters. This study shed light on the effect of pump design on the performance and risk of blood damage, indicating the roles of the hub shape and angular velocity as dominant parameters.

Impact of rise and fall phases of shear on platelet activation and aggregation using microfluidics.

Blood flow disorders are often the result of the non-physiological narrowing of blood arteries caused by atherosclerosis and thrombus. The blood then proceeds through rising-peak-decreasing phases as it passes through the narrow area. Although abnormally high shear is known to activate platelets, the shear process that platelets undergo in small arteries is complex. Thus, understanding how each shear phase affects platelet activation can be used to improve antiplatelet therapy and decrease the risk of side effects like bleeding. Blood samples were sheared (68.8 ms,5200s-1) in vitro by the microfluidic technique, and platelet activation levels (P-selectin and integrin αIIbβ3) and von Willebrand factor (vWF) binding to platelets were analyzed by flow cytometry. Post-stenosis platelet aggregation was dynamically detected using microfluidic technology. We studied TXA2, P2Y12-ADP, and integrin αIIbβ3-fibrinogen receptor pathways by adding antiplatelet drugs, such as acetylsalicylic acid (ASA, an active ingredient of aspirin that inhibits platelet metabolism), ticagrelor (hinders platelet activation), and tirofiban (blocks integrin αIIbβ3 receptor) in vitro, respectively, to determine platelet activation function mediated by transient non-physiological high shear rates. We demonstrated that platelets can be activated under transient pathological high shear rates. The shear rise and fall phases influenced shear-induced platelet activation by regulating the binding of vWF to platelets. The degree of platelet activation and aggregation increased with multiple shear rise and fall phases. ASA did not inhibit shear-mediated platelet activation, but ticagrelor and tirofiban effectively inhibited shear-mediated platelet activation. Our data demonstrated that the shear rise and fall phases play an important role in shear-mediated platelet activation and promote platelet activation and aggregation in a vWF-dependent manner. Blocking integrin αIIbβ3 receptor and hindering P2Y12-ADP were beneficial to reducing shear-mediated platelet activation.

Investigation of the role of von Willebrand factor in shear-induced platelet activation and functional alteration under high non-physiological shear stress.

von Willebrand factor (vWF) plays a crucial role in physiological hemostasis through platelet and subendothelial collagen adhesion. However, its role in shear-induced platelet activation and functional alteration under non-physiological conditions common to blood-contacting medical devices (BCMDs) is not well investigated. Fresh healthy human blood was treated with an anti-vWF antibody to block vWF-GPIbα interaction. Untreated blood was used as a control. They were exposed to three levels of non-physiological shear stress (NPSS) (75, 125, and 175 Pa) through a shearing device with an exposure time of 0.5 s to mimic typical shear conditions in BCMDs. Flow cytometric assays were used to measure the expression levels of PAC-1 and P-Selectin and platelet aggregates for platelet activation and the expression levels of GPIbα, GPIIb/IIIa, and GPVI for receptor shedding. Collagen/ristocetin-induced platelet aggregation capacity was characterized by aggregometry. The levels of platelet activation and aggregates increased with increasing NPSS in the untreated blood. More receptors were lost with increasing NPSS, resulting in a decreased capacity of collagen/ristocetin-induced platelet aggregation. In contrast, the increase in platelet activation and aggregates after exposure to NPSS, even at the highest level of NPSS, was significantly lower in treated blood. Nevertheless, there was no notable difference in receptor shedding, especially for GPIIb/IIIa and GPVI, between the two blood groups at the same level of NPSS. The block of vWF exacerbated the decreased capacity of collagen/ristocetin-induced platelet aggregation. High NPSS activates platelets mainly by enhancing the vWF-GPIbα interaction. Platelet activation and receptor shedding induced by high NPSS likely occur through different pathways.

Cordycepin Inhibits Pathological High Shear-Induced Platelet Aggregation, Activation, and Phosphatidylserine Exposure by Regulating the Phosphorylation of Akt Protein

Cordycepin Inhibits Pathological High Shear-Induced Platelet Aggregation, Activation, and Phosphatidylserine Exposure by Regulating the Phosphorylation of Akt Protein

Experimental Investigation of Microfluidic Device for Platelet Activation.

Wound healing is accelerated when Platelet Rich Plasma is activated and growth factors are released. In this study, it was aimed to stimulate platelets without using chemical stimulants. Two types of mechanical platelet activation methods have been proposed in this study. The first one is a microfluidic chip developed with the shear-induced platelet activation approach. The second one is a piezo-based ultrasound-assisted device which provides platelet activation by stimulating with an ultrasonic wave (0.55 and 1.1 MHz). Three different microfluidic chip designs were worked out to determine the optimal shear stress characteristics; 8-nodes (2789 µs, 288 shear pulses, and 98.3 dyne/cm2), 40-nodes (2765 µs, 1440 shear pulses, and 95.5 dyne/cm2) and pillar-shaped (1030 µs, 1656 shear pulses, and 48.1 dyne/cm2). The highest platelet activation rate (72.7%) was obtained from the chips with 8-nodes. In the ultrasound-assisted device, 32.4% activation rate was obtained from ultrasound waves with 0.55 MHz frequency and 10 Vp-p amplitude. These activation rates, determined by CD62P (P-Selectin) expression, are significantly higher than spontaneous activation of intact platelets (8.5%). In addition, the gradual increase in activation of stimulated platelets with incubation at room temperature showed that activation continued after stimulation. The results showed that these microfluidic devices can be used for platelet activation to enhance the effect of PRP treatment and might reduce adverse immune reactions that may happened due to the use of exogenous activator substances. Fast-response, low-cost, easy-to-use and controllable biomedical device have been developed for PRP applications.

Patient specific approach to analysis of shear-induced platelet activation in haemodialysis arteriovenous fistula.

Shear-induced platelet activation (SIPAct) is an important mechanism of thrombosis initiation under high blood flow. This mechanism relies on the interaction of platelets with the von Willebrand factor (VWF) capable of unfolding under high shear stress. High shear stress occurs in the arteriovenous fistula (AVF) commonly used for haemodialysis. A novel patient-specific approach for the modelling of SIPAct in the AVF was proposed. This enabled us to estimate the SIPAct level via computational fluid dynamics. The suggested approach was applied for the SIPAct analysis in AVF geometries reconstructed from medical images. The approach facilitates the determination of the SIPAct level dependence on both biomechanical (AVF flow rate) and biochemical factors (VWF multimer size). It was found that the dependence of the SIPAct level on the AVF flow rate can be approximated by a power law. The critical flow rate was a decreasing function of the VWF multimer size. Moreover, the critical AVF flow rate highly depended on patient-specific factors, e.g., the vessel geometry. This indicates that the approach may be adopted to elucidate patient-specific thrombosis risk factors in haemodialysis patients.

Investigation of shear-induced platelet activation in ventricular assist device

Due to the scarcity of organ donations, ventricular assist devices are the most accessible treatment for patients with advanced heart failure. Since their development, these devices have helped thousands of patients and could have helped even more had it not been for some of the complications they still experience. Among the most common complications are thrombosis and hemolysis. The purpose of this paper is to numerically investigate the effect of the blade angle and the blade count on the hydraulic properties of a newly designed ventricular assist device, as well as the potential for shear-induced platelet activation. The study was conducted on several models with different blade angles and blade counts using a variety of rotational speeds. Analysis of the obtained results showed a significant improvement in the pressure rise and the hydraulic efficiency in models with higher blade angle and lower blade count. In contrast, the other models showed slight improvement or deterioration of the hydraulic performance. In terms of shear-induced platelet activation, although the performance of all models was within an acceptable range, models with a higher blade angle and lower blade count had the lowest average platelet activation state.

Computational Analysis of Flow Structures in Turbulent Ventricular Blood Flow Associated With Mitral Valve Intervention.

Cardiac disease and clinical intervention may both lead to an increased risk for thrombosis events due to a modified blood flow in the heart, and thereby a change in the mechanical stimuli of blood cells passing through the chambers of the heart. Specifically, the degree of platelet activation is influenced by the level and type of mechanical stresses in the blood flow. In this article we analyze the blood flow in the left ventricle of the heart through a computational model constructed from patient-specific data. The blood flow in the ventricle is modelled by the Navier-Stokes equations, and the flow through the mitral valve by a parameterized model which represents the projected opening of the valve. A finite element method is used to solve the equations, from which a simulation of the velocity and pressure of the blood flow is constructed. The intraventricular blood flow is complex, in particular in diastole when the inflow jet from the atrium breaks down into turbulent flow on a range of scales. A triple decomposition of the velocity gradient tensor is then used to distinguish between rigid body rotational flow, irrotational straining flow, and shear flow. The triple decomposition enables the separation of three fundamentally different flow structures, that each generates a distinct type of mechanical stimulus on the blood cells in the flow. We compare the results in a simulation where a mitral valve clip intervention is modelled, which leads to a significant modification of the intraventricular flow. Further, we perform a sensitivity study of the results with respect to the positioning of the clip. It was found that the shear in the simulation cases treated with clips increased more compared to the untreated case than the rotation and strain did. A decrease in valve opening area of 64% in one of the cases led to a 90% increase in rotation and strain, but a 150% increase in shear. The computational analysis opens up for improvements in models of shear-induced platelet activation, by offering an algorithm to distinguish shear from other modalities in intraventricular blood flow.

Continuum modeling of thrombus formation and growth under different shear rates

Obstruction of blood flow due to thrombosis is a major cause of ischemic stroke, myocardial infarction, and in severe cases, mortality. In particular, in blood wetting medical devices, thrombosis is a common reason for failure. The prediction of thrombosis by understanding signaling pathways using computational models, lead to identify the risk of thrombus formation in blood-contacting devices and design improvements. In this study, a mathematical model of thrombus formation and growth is presented. A biochemical model of platelet activation and aggregation is developed to predict thrombus size and shape at the site of vascular injury. Computational fluid dynamics using the finite volume method is employed to compute the velocity and pressure fields which are influenced by the growing thrombi. The passive transport of platelets, agonists, the platelet activation kinetics, their adhesion to the growing thrombi and embolization of platelets are solved by a fully coupled set of convection–diffusion–reaction equations. The thrombogenic surface representing blood-contacting material or injured blood vessel was incorporated into the model as a surface flux boundary condition to initiate thrombus formation. The blood is considered as a Newtonian fluid, while the thrombus is treated as a porous medium. The results are compared with in vitro experiments of a microfluidic chamber at an initial inlet venous shear rate of 200s−1 using a pressure–inlet boundary condition. The thrombus development due to agonist concentrations and change in the shear rate as well as thromboembolism for this benchmark problem is successfully computed. Furthermore, to extend the current model to a physiologically relevant configuration, thrombus formation in a blood tube is simulated. Two different heterogeneous reaction rates for platelet aggregation are used to simulate thrombus growth under a constant inlet flow rate. The findings show that the thrombus shape can be substantially altered by the hemodynamic conditions, increase in the shear rate and due to the combined effects of shear induced platelet activation and the heterogeneous reaction rates. It is also concluded that the model is able to predict thrombus formation in different physiological and pathological hemodynamics.

Investigation of the Role of Von Willebrand Factor in Shear-Induced Platelet Activation and Functional Alternation Under High Non-Physiological Shear Stress

Investigation of the Role of Von Willebrand Factor in Shear-Induced Platelet Activation and Functional Alternation Under High Non-Physiological Shear Stress

Models of Shear-Induced Platelet Activation and Numerical Implementation With Computational Fluid Dynamics Approaches.

Shear-induced platelet activation is one of the critical outcomes when blood is exposed to elevated shear stress. Excessively activated platelets in the circulation can lead to thrombus formation and platelet consumption, resulting in serious adverse events such as thromboembolism and bleeding. While experimental observations reveal that it is related to the shear stress level and exposure time, the underlying mechanism of shear-induced platelet activation is not fully understood. Various models have been proposed to relate shear stress levels to platelet activation, yet most are modified from the empirically calibrated power-law model. Newly developed multiscale platelet models are tested as a promising approach to capture a single platelet's dynamic shape during activation, but it would be computationally expensive to employ it for a large-scale analysis. This paper summarizes the current numerical models used to study the shear-induced platelet activation and their computational applications in the risk assessment of a particular flow pattern and clot formation prediction.

Mechanobiology of shear-induced platelet aggregation leading to occlusive arterial thrombosis: A multiscale in silico analysis

Occlusive thrombosis in arteries causes heart attacks and strokes. The rapid growth of thrombus at elevated shear rates (~10,000 1/s) relies on shear-induced platelet aggregation (SIPA) thought to come about from the entanglement of von Willebrand factor (VWF) molecules. The mechanism for SIPA is not yet understood in terms of cell- and molecule-level dynamics in fast flowing bloodstreams. Towards this end, we develop a multiscale computational model to recreate SIPA in silico, where the suspension dynamics and interactions of individual platelets and VWF multimers are resolved directly. The platelet-VWF interaction via GP1b-A1 bonds is prescribed with intrinsic binding rates theoretically derived and informed by single-molecule measurements. The model is validated against existing microfluidic SIPA experiments, showing good agreement with the in vitro observations in terms of the morphology, traveling distance and capture time of the platelet aggregates. Particularly, the capture of aggregates can occur in a few milliseconds, comparable to the platelet transit time through pathologic arterial stenotic sections and much shorter than the time for shear-induced platelet activation. The multiscale SIPA simulator provides a cross-scale tool for exploring the biophysical mechanisms of SIPA in silico that are difficult to access with single-molecule measurements or micro-/macro-fluidic assays only.

Microfluidic assay of antiplatelet agents for inhibition of shear-induced platelet adhesion and activation.

We have developed a microfluidic system to perfuse whole blood through a flow channel with an upstream stenotic region and a downstream protein capture region. This flow-based system was used to assay how effectively antiplatelet agents suppress shear-induced platelet adhesion and activation downstream of the stenotic region. Microcontact printing was used to covalently attach one of three platelet binding proteins [fibrinogen, collagen, or von Willebrand factor (vWf)] to the surface of the downstream capture region. Whole blood with an antiplatelet agent was transiently exposed to an upstream high wall shear rate (either 4860 s-1 or 11 560 s-1), and subsequently flowed over the downstream capture region where the platelet adhesion was measured. Several antiplatelet agents (acetylsalicylic acid, tirofiban, eptifibatide, anti-vWf, and anti-GPIbα) were evaluated for their efficacy in attenuating downstream adhesion. Following antibody blocking of vWf or GPIbα, downstream platelet activation was also assessed in perfused blood by flow cytometry using two activation markers (active GPIIb/IIIa and P-selectin). Acetylsalicylic acid demonstrated its inability to diminish shear-induced platelet adhesion to all three binding proteins. GPIIb/IIIa inhibitors (tirofiban and eptifibatide) significantly reduced platelet adhesion to fibrinogen. Antibody blocking of vWf or GPIbα effectively diminished platelet adhesion to all three capture proteins as well as platelet activation in perfused blood, indicating an essential role of vWf-GPIbα interaction in mediating shear-induced platelet aggregation.

Shear-Induced Platelet Activation is Sensitive to Age and Calcium Availability: A Comparison of Adult and Cord Blood.

Antiplatelet therapy for neonates and infants is often extrapolated from the adult experience, based on limited observation of agonist-induced neonatal platelet hypoactivity and poor understanding of flow shear-mediated platelet activation. Therefore, thrombotic events due to device-associated disturbed flow are inadequately mitigated in critically ill neonates with indwelling umbilical catheters and infants receiving cardiovascular implants. Whole blood (WB), platelet-rich plasma (PRP), and gel-filtered platelets (GFP) were prepared from umbilical cord and adult blood, and exposed to biochemical agonists or pathological shear stress of 70 dyne/cm2. We evaluated α-granule release, phosphatidylserine (PS) scrambling, and procoagulant response using P-selectin expression, Annexin V binding, and thrombin generation (PAS), respectively. Activation modulation due to depletion of intracellular and extracellular calcium, requisite second messengers, was also examined. Similar P-selectin expression was observed for sheared adult and cord platelets, with concordant inhibition due to intracellular and extracellular calcium depletion. Sheared cord platelet Annexin V binding and PAS activity was similar to adult values in GFP, but lower in PRP and WB. Annexin V on sheared cord platelets was calcium-independent, with PAS slightly reduced by intracellular calcium depletion. Increased PS activity on purified sheared cord platelets suggest that their intrinsic function under pathological flow conditions is suppressed by cell-cell or plasmatic components. Although secretory functions of adult and cord platelets retain comparable calcium-dependence, PS exposure in sheared cord platelets is uniquely calcium-independent and distinct from adults. Identification of calcium-regulated developmental disparities in shear-mediated platelet function may provide novel targets for age-specific antiplatelet therapy.

Platelet activation via dynamic conformational changes of von Willebrand factor under shear.

Shear-induced conformational changes of von Willebrand factor (VWF) play an important role in platelet activation. A novel approach describing VWF unfolding on the platelet surface under dynamic shear stress is proposed. Cumulative effect of dynamic shear on platelet activation via conformational changes of VWF is analysed. The critical condition of shear-induced platelet activation is formulated. The explicit expression for the threshold value of cumulative shear stress as a function of VWF multimer size is derived. The results open novel prospects for pharmacological regulation of shear-induced platelet activation through control of VWF multimers size distribution.

Shear-Dependent Platelet Aggregation: Mechanisms and Therapeutic Opportunities.

Cardiovascular diseases (CVD) are the number one cause of morbidity and death worldwide. As estimated by the WHO, the global death rate from CVD is 31% wherein, a staggering 85% results from stroke and myocardial infarction. Platelets, one of the key components of thrombi, have been well-investigated over decades for their pivotal role in thrombus development in healthy as well as diseased blood vessels. In hemostasis, when a vascular injury occurs, circulating platelets are arrested at the site of damage, where they are activated and aggregate to form hemostatic thrombi, thus preventing further bleeding. However, in thrombosis, pathological activation of platelets occurs, leading to uncontrolled growth of a thrombus, which in turn can occlude the blood vessel or embolize, causing downstream ischemic events. The molecular processes causing pathological thrombus development are in large similar to the processes controlling physiological thrombus formation. The biggest challenge of anti-thrombotics and anti-platelet therapeutics has been to decouple the pathological platelet response from the physiological one. Currently, marketed anti-platelet drugs are associated with major bleeding complications for this exact reason; they are not effective in targeting pathological thrombi without interfering with normal hemostasis. Recent studies have emphasized the importance of shear forces generated from blood flow, that primarily drive platelet activation and aggregation in thrombosis. Local shear stresses in obstructed blood vessels can be higher by up to two orders of magnitude as compared to healthy vessels. Leveraging abnormal shear forces in the thrombus microenvironment may allow to differentiate between thrombosis and hemostasis and develop shear-selective anti-platelet therapies. In this review, we discuss the influence of shear forces on thrombosis and the underlying mechanisms of shear-induced platelet activation. Later, we summarize the therapeutic approaches to target shear-sensitive platelet activation and pathological thrombus growth, with a particular focus on the shear-sensitive protein von Willebrand Factor (VWF). Inhibition of shear-specific platelet aggregation and targeted drug delivery may prove to be much safer and efficacious approaches over current state-of-the-art antithrombotic drugs in the treatment of cardiovascular diseases.

Evaluation of in vitro hemolysis and platelet activation of a newly developed maglev LVAD and two clinically used LVADs with human blood.

In vitro hemolysis testing remains one of the most important performance measures to judge the hemocompatibility of a left ventricular assist device (LVAD). Clinically relevant operating conditions and appropriate testing blood are essential to infer in vitro data for potential clinical use. This in vitro study was carried out to evaluate and compare the hemolytic performance of a newly developed magnetically levitated (maglev) LVAD (CH-VAD) with two clinically used LVADs (HVAD and HeartMate II (HMII)) using fresh human blood. A small volume (~300 mL) in vitro circulating flow loop was constructed with a LVAD generated flow of 4.5 L/min at the nominal or reported clinical operating speed for each LVAD. The blood was circulated in the loop for 4 hours with samples drawn at baseline and hourly. Plasma-free hemoglobin (PFH) concentrations in the hourly blood samples were determined with spectrophotometry. Normalized index of hemolysis (NIH) was calculated to compare the hemolytic performance of the CH-VAD and the two reference LVADs. Platelet activation was measured with flow cytometry. The experimental test for each device was repeated at least 7 times. The data from this study showed that all the three LVADs generated very low hemolysis (NIH <0.01 g/100L). The CH-VAD was found to have a significantly lower NIH value (0.00135 ± 0.00032 g/100L) compared to the HVAD (0.00525 ± 0.00183 g/100L) and the HMII (0.00583 ± 0.00182 g/100L). No statistically significant difference in device-generated hemolysis was found between the HVAD and the HMII. The level of platelet activation induced by the CH-VAD is significantly lower than those by the HVAD and the HMII. The data suggest that the shear-induced hemolysis and platelet activation of the CH-VAD are acceptable relative to the two LVADs currently in clinical use.

Shear-induced platelet adherence and activation in an in-vitro dynamic multiwell-plate system.

Circulating blood cells are prone to varying flow conditions when contacting cardiovascular devices. For a profound understanding of the complex interplay between the blood components/cells and cardiovascular implant surfaces, testing under varying shear conditions is required. Here, we study the influence of arterial and venous shear conditions on the in vitro evaluation of the thrombogenicity of polymer-based implant materials.Medical grade poly(dimethyl siloxane) (PDMS), polyethylene terephthalate (PET) and polytetrafluoroethylene (PTFE) films were included as reference materials. The polymers were exposed to whole blood from healthy humans. Blood was agitated orbitally at low (venous shear stress: 2.8 dyne · cm-2) and high (arterial shear stress: 22.2 dyne · cm-2) agitation speeds in a well-plate based test system. Numbers of non-adherent platelets, platelet activation (P-Selectin positive platelets), platelet function (PFA100 closure times) and platelet adhesion (laser scanning microscopy (LSM)) were determined.Microscopic data and counting of the circulating cells revealed increasing numbers of material-surface adherent platelets with increasing agitation speed. Also, activation of the platelets was substantially increased when tested under the high shear conditions (P-Selectin levels, PFA-100 closure times). At low agitation speed, the platelet densities did not differ between the three materials. Tested at the high agitation speed, lowest platelet densities were observed on PDMS, intermediate levels on PET and highest on PTFE. While activation of the circulating platelets was affected by the implant surfaces in a similar manner, PFA closure times did not reflect this trend.Differences in the thrombogenicity of the studied polymers were more pronounced when tested at high agitation speed due to the induced shear stresses. Testing under varying shear stresses, thus, led to a different evaluation of the implant thrombogenicity, which emphasizes the need for testing under various flow conditions. Our data further confirmed earlier findings where the same reference implants were tested under static (and not dynamic) conditions and with fresh human platelet rich plasma instead of whole blood. This supports that the application of common reference materials may improve inter-study comparisons, even under varying test conditions.

Fluid Dynamic Characterization of Transcatheter Aortic Valves Using Particle Image Velocimetry.

Transcatheter aortic valves provide superior systolic hemodynamic performance in terms of valvular pressure gradient and effective orifice area compared with equivalent size surgical bioprostheses. However, in depth investigation of the flow field structures is of interest to examine the flow field characteristics and provide experimental evidence necessary for validation of computational models. The goal of this study was to compare flow field characteristics of the three most commonly used transcatheter and surgical valves using phase-locked particle image velocimetry (PIV). 26-mm Edwards SAPIEN 3, 26-mm Medtronic CoreValve, and 25-mm Carpentier-Edwards PERIMOUNT Magna were examined in a pulse duplicator with input parameters matching ISO-5840, that is, heart rate of 70 beats/min, cardiac output of 5 L/min, and mean aortic pressure of 100 mm Hg. A 2D PIV system was used to obtain flow velocity and viscous shear stress fields during the entire cardiac cycle. In vitro testing showed that the mean transvalvular pressure gradient was lowest for SAPIEN 3, followed by CoreValve, and PERIMOUNT Magna surgical bioprosthesis. In addition, the viscous shear stress magnitude within the jet boundary layer was higher in PERIMOUNT Magna than CoreValve and SAPIEN 3 at the peak of the flow. However, the measured shear stress values were below the known threshold for platelet activation and red blood damage. Therefore, shear-induced platelet activation is unlikely to take place during systole in the three bioprosthetic heart valves. The PIV measurements can be used for verification and validation of computational simulations.

The physical spacing between the von Willebrand factor D'D3 and A1 domains regulates platelet adhesion in vitro and in vivo

Background Previous exvivo studies using truncated VWF (von Willebrand factor) suggest that domain-level molecular architecture may control platelet-GpIbα binding function. Objective We determined if this is the case with multimeric VWF invivo. Methods Full-length human VWF ('hV') was modified with a 22-amino acid mucinous stretch at either the N-terminus of VWF-A1 to create 'hNV' or C-terminus to yield 'hCV'. This extends the physical distance between VWF-A1 and the adjacent domains by ~6nm. Similar mucin inserts were also introduced into a human-murine chimera ('h[mA1]V') where murine-A1 replaced human-A1 in hV. This yielded 'h[mA1]NV' and 'h[mA1]CV', with N- and C-terminal inserts. The constructs were tested exvivo and invivo. Results Mucin insertion at the N-terminus, but not C-terminus, in both types of constructs resulted in >50-fold increase in binding to immobilized GpIbα. N-terminal insertion also resulted in greater shear-induced platelet activation, more thrombus formation on collagen, enhanced platelet accumulation and slower platelet translocation on immobilized VWF in microfluidics assays. Hydrodynamic injection-based expression of h[mA1]NV, but not h[mA1]V or h[mA1]CV, in VWF-/- mice caused profound thrombocytopenia, reduced plasma VWF concentrations, lower multimer distribution, and incessant tail bleeding that is reminiscent of von Willebrand disease type 2B. Platelet plugs were noted in the portal veins and hepatic arteries. An anti-D'D3 mAb DD3.3 that displays enhanced binding to VWF containing the N-terminal mucin insert also exhibited increased binding to wild-type VWF under shear and upon ristocetin addition. Conclusion Conformation changes at the VWF D'D3-A1 interface may be a key regulator of thrombosis invivo. Structural features at the A1-A2 interface are likely of less significance.