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Background It has been demonstrated that von Willebrand factor (VWF) mediated platelet-endothelium and platelet-platelet interactions are shear dependent. The VWF’s mobility under dynamic conditions (e.g. flow) is pivotal to platelet adhesion and VWF-mediated aggregate formation in the cascade of VWF-platelet interactions in haemostasis. Results Combining microfluidic tools with fluorescence and reflection interference contrast microscopy (RICM), here we show, that specific deletions in the A-domains of the biopolymer VWF affect both, adhesion and aggregation properties independently. Intuitively, the deletion of the A1-domain led to a significant decrease in both adhesion and aggregate formation of platelets. Nevertheless, the deletion of the A2-domain revealed a completely different picture, with a significant increase in formation of rolling aggregates (gain of function). We predict that the A2-domain effectively ‘masks’ the potential between the platelet glycoprotein (GP) Ib and the VWF A1-domain. Furthermore, the deletion of the A3-domain led to no significant variation in either of the two functional characteristics. Conclusions These data demonstrate that the macroscopic functional properties i.e. adhesion and aggregate formation cannot simply be assigned to the properties of one particular domain, but have to be explained by cooperative phenomena. The absence or presence of molecular entities likewise affects the properties (thermodynamic phenomenology) of its neighbours, therefore altering the macromolecular function.

Platelet adhesion and activation are essential initial processes of arterial and microvascular hemostasis, where high hydrodynamic forces from the bloodflow impede coagulation. The process relies on von Willebrand factor (VWF)-a linear multimeric protein of blood plasma plays a pivotal role in mechanochemical regulation of shear-induced platelet aggregation (SIPA). Adhesive interactions between VWF and glycoprotein receptors GPIb are crucial for platelet recruitment under high shear stress in fluid. Recent advances in experimental studies revealed that mechanical tension on the extracellular part of GPIb may trigger a cascade of biochemical reactions in platelets leading to activation of integrins [Formula: see text] (also known as GPIIb/IIIa) and strengthening of the adhesion. The present paper is aimed at investigation of this process by three-dimensional computer simulations of platelet adhesion to surface-grafted VWF multimers in pressure-driven flow of platelet-rich plasma. The simulations demonstrate that GPIb-mediated mechanotransduction is a feasible way of platelet activation and stabilization of platelet aggregates under high shear stress. Quantitative understanding of mechanochemical processes involved in SIPA would potentially promote the discovery of new anti-platelet medication and the development of multiscale numerical models of platelet thrombosis and hemostasis.

The purpose of this study was to investigate the influence of non-physiological high shear stress on activation and shedding of platelet GP IIb/IIIa receptors. The healthy donor blood was exposed to three levels of high shear stresses (25, 75, 125 Pa) from the physiological to non-physiological status with three short exposure time (0.05, 0.5, 1.5 s), created by a specific blood shearing system. The activation and shedding of the platelet GPIIb/IIIa were analyzed using flow cytometry and enzyme-linked immunosorbent assay. In addition, platelet P-selectin expression of sheared blood, which is a marker for activated platelets, was also analyzed. The results from the present study showed that the number of activated platelets, as indicated by the surface GPIIb/IIIa activation and P-selectin expression, increased with increasing the shear stress level and exposure time. However, the mean fluorescence of GPIIb/IIIa on the platelet surface, decreased with increasing the shear stress level and exposure time. The reduction of GPIIb/IIIa on the platelet surface was further proved by the reduction of further activated platelet GPIIb/IIIa surface expression induced by ADP and the increase in GPIIb/IIIa concentration in microparticle-free plasma with increasing the applied shear stress and exposure time. It is clear that non-physiological shear stress induce a paradoxical phenomenon, in which both activation and shedding of the GPIIb/IIIa on the platelet surface occur simultaneously. This study may offer a new perspective to explain the reason of both increased thrombosis and bleeding events in patients implanted with high shear blood-contacting medical devices.

We report a novel device to analyze cell-surface interactions under controlled fluid-shear conditions on well-characterised protein surfaces. Its performance is demonstrated by studying platelets interacting with immobilised von Willebrand Factor at arterial vascular shear rates using just 200 μL of whole human blood per assay. The device's parallel-plate flow chamber, with 0.1 mm² cross sectional area and height-to-width ratio of 1:40, provides uniform, well-defined shear rates along the chip surface with negligible vertical wall effects on the fluid flow profile while minimizing sample volumetric flow. A coating process was demonstrated by ellipsometry, atomic force microscopy, and fluorescent immunostaining to provide reproducible, homogeneous, uniform protein layers over the 0.7 cm² cell-surface interaction area. Customized image processing quantifies dynamic cellular surface coverage vs. time throughout the whole-blood-flow assay for a given drug treatment or disease state. This device can track the dose response of anti-platelet drugs, is suitable for point-of-care diagnostics, and is designed for adaptation to mass manufacture.

Shear-mediated platelet activation (SMPA) in the “free flow” is the net result of a range of cell mechanobiological mechanisms. Previously, we outlined three main groups of mechanisms including: 1) mechano-destruction - i.e. additive platelet (membrane) damage; 2) mechano-activation - i.e. activation of shear-sensitive ion channels and pores; and 3) mechano-transduction - i.e. “outside-in” signaling via a range of transducers. Here, we report on recent advances since our original report which describes additional features of SMPA. A clear “signature” of SMPA has been defined, allowing differentiation from biochemically-mediated activation. Notably, SMPA is characterized by mitochondrial dysfunction, platelet membrane eversion, externalization of anionic phospholipids, and increased thrombin generation on the platelet surface. However, SMPA does not lead to integrin αIIbβ3 activation or P-selectin exposure due to platelet degranulation, as is commonly observed in biochemical activation. Rather, downregulation of GPIb, αIIbβ3, and P-selectin surface expression is evident. Furthermore, SMPA is accompanied by a decrease in overall platelet size coupled with a concomitant, progressive increase in microparticle generation. Shear-ejected microparticles are highly enriched in GPIb and αIIbβ3. These observations indicate the enhanced diffusion, migration, or otherwise dispersion of platelet adhesion receptors to membrane zones, which are ultimately shed as receptor-rich PDMPs. The pathophysiological consequence of this progressive shear accumulation phenomenon is an associated dyscrasia of remaining platelets – being both reduced in size and less activatable via biochemical means – a tendency to favor bleeding, while concomitantly shed microparticles are highly prothrombotic and increase the tendency for thrombosis in both local and systemic milieu. These mechanisms and observations offer direct clinical utility in allowing measurement and guidance of the net balance of platelet driven events in patients with implanted cardiovascular therapeutic devices.

The paper presents the results of a study of the effects of separation of spherical particles with different densities in a fast gravitational flow on a rough slope with varying flow parameters in order to increase the efficiency of the process. The determining role of the effect of quasi-diffusion separation in the process has been established, the intensive manifestation of which is facilitated by the formation of s-shaped profiles of the fraction of voids and velocity in the flow. The results of an experimental and analytical study of the efficiency of particle separation by density in a fast gravitational flow when shear deformations are activated by longitudinal pulses are presented. It has been established that when particles are exposed to the pulse effect from the open surface of the flow, its central part develops a zone with a typical s-shaped inflection in the velocity profile and void fraction, which functions like a quasi-diffusion separator that increases the efficiency of the process.

Individuals with mechanical heart valve implants are plagued by flow-induced thromboembolic complications, which are undoubtedly caused by platelet activation. Flow fields in or around the affected regions involve brief exposure to pathologically high-shear stresses on the order of 100 to 1000 dyne/cm². Although high shear is known to activate platelets directly, their subsequent behavior is not known. We hypothesize that the post-high-shear activation behavior of platelets is particularly relevant in understanding the increased thrombotic risk associated with blood-recirculating prosthetic cardiovascular devices. Purified platelets were exposed to brief (5-40 s) periods of high-shear stress, and then exposed to longer periods (15-60 min) of low shear. Their activation state was measured using a prothrombinase-based assay. Platelets briefly exposed to an initial high-shear stress (e.g., 60 dyne/cm² for 40 s) activate a little, but this study shows that they are now sensitized, and when exposed to subsequent low shear stress, they activate at least 20-fold faster than platelets not initially exposed to high shear. The results show that platelets in vitro exposed beyond a threshold of high-shear stress are primed for subsequent activation under normal cardiovascular circulation conditions, and they do not recover from the initial high-shear insult.

In the United States, over 125,000 mechanical heart valves (MHVs) are implanted each year. Flow through the MHV hinge can cause thromboemboli formation. The purpose of this study was to examine various orifice geometries representing the MHV hinge region and how these geometries may contribute to platelet activation and thrombin generation. We also characterized these flow fields with digital particle image velocimetry (DPIV). Citrated human blood at room temperature was forced through the orifices (400 and 800 μm ID) with a centrifugal bypass pump, continuously infusing calcium chloride to partially reverse the citrate anticoagulant. Blood samples were tested for the presence of thrombin–antithrombin complex (TAT) and platelet factor 4 (PF4). Velocity and shear stress were measured with DPIV using a blood analog fluid seeded with fluorescent microbeads. The results indicate that small changes in geometry, although they do not affect the bulk flow, change the coagulation propensity as blood flows through the orifices. A more abrupt geometry allows more stagnation to occur resulting in more thrombin generation. PF4 measurements indicated similar levels of platelet activation for all orifices. DPIV showed differences in the jets with respect to entrainment of stagnant fluid. These results help to pinpoint the important parameters that lead to flow stasis and subsequent thrombus formation.

Thrombosis is a major contributor to cardiovascular disease, which can lead to myocardial infarction and stroke. Thrombosis may form in tortuous microvessels, which are often seen throughout the human body, but the microscale mechanisms and processes are not well understood. In straight vessels, the presence of red blood cells (RBCs) is known to push platelets toward walls, which may affect platelet aggregation and thrombus formation. However in tortuous vessels, the effects of RBC interactions with platelets in thrombosis are largely unknown. Accordingly, the objective of this work was to determine the physical effects of RBCs, platelet size, and vessel tortuosity on platelet activation and thrombus formation in tortuous arterioles. A discrete element computational model was used to simulate the transport, collision, adhesion, aggregation, and shear-induced platelet activation of hundreds of individual platelets and RBCs in thrombus formation in tortuous arterioles. Results showed that high shear stress near the inner sides of curved arteriole walls activated platelets to initiate thrombosis. RBCs initially promoted platelet activation, but then collisions of RBCs with mural thrombi reduced the amount of mural thrombus and the size of emboli. In the absence of RBCs, mural thrombus mass was smaller in a highly tortuous arteriole compared to a less tortuous arteriole. In the presence of RBCs however, mural thrombus mass was larger in the highly tortuous arteriole compared to the less tortuous arteriole. As well, smaller platelet size yielded less mural thrombus mass and smaller emboli, either with or without RBCs. This study shed light on microscopic interactions of RBCs and platelets in tortuous microvessels, which have implications in various pathologies associated with thrombosis and bleeding.

Shear thickening in dense particulate suspensions was recently proposed to be driven by the activation of friction above an onset stress needed to overcome repulsive forces between particles. Testing this scenario represents a major challenge because classical rheological approaches do not provide access to the frictional properties of suspensions. Here we adopt a different strategy inspired by pressure-imposed configurations in granular flows that specifically gives access to this information. By investigating the quasi-static avalanche angle, compaction, and dilatancy effects in different nonbuoyant suspensions flowing under gravity, we demonstrate that particles in shear-thickening suspensions are frictionless under low confining pressure. Moreover, we show that tuning the range of the repulsive force below the particle roughness suppresses the frictionless state and also the shear-thickening behavior of the suspension. These results, which link microscopic contact physics to the suspension macroscopic rheology, provide direct evidence that the recent frictional transition scenario applies in real suspensions.