78 research outputs found
A membrane-based microfluidic device for controlling the flux of platelet agonists into flowing blood
The flux of platelet agonists into flowing blood is a critical event in thrombosis and hemostasis. However, few in vitro methods exist for examining and controlling the role of platelet agonists on clot formation and stability under hemodynamic conditions. In this paper, we describe a membrane-based method for introducing a solute into flowing blood at a defined flux. The device consisted of a track-etched polycarbonate membrane reversibly sealed between two microfluidic channels; one channel contained blood flowing at a physiologically relevant shear rate, and the other channel contained the agonist(s). An analytical model described the solute flux as a function of the membrane permeability and transmembrane pressure. The model was validated using luciferase as a model solute for transmembrane pressures of 50–400 Pa. As a proof-of-concept, the weak platelet agonist ADP was introduced into whole blood flowing at 250 s-1 at three fluxes (1.5, 2.4, and 4.4 × 10-18 mol µm-2 s-1). Platelet aggregation was monitored by fluorescence microscopy during the experiment and the morphology of aggregates was determined by post hoc confocal and electron microscopy. At the lowest flux (1.5 × 10-18 mol µm-2 s-1), we observed little to no aggregation. At the higher fluxes, we observed monolayer (2.4 × 10-18 mol µm-2 s-1) and multilayer (4.4 × 10-18 mol µm-2 s-1) aggregates of platelets and found that the platelet density within an aggregate increased with increasing ADP flux. We expect this device to be a useful tool in unraveling the role of platelet agonists on clot formation and stability
Isotopically nonstationary 13C metabolic flux analysis in resting and activated human platelets
Platelet metabolism is linked to platelet hyper- and hypoactivity in numerous human diseases. Developing a detailed understanding of the link between metabolic shifts and platelet activation state is integral to improving human health. Here, we show the first application of isotopically nonstationar
Zoster-Associated Prothrombotic Plasma Exosomes and Increased Stroke Risk
Herpes zoster (HZ; shingles) caused by varicella zoster virus reactivation increases stroke risk for up to 1 year after HZ. The underlying mechanisms are unclear, however, the development of stroke distant from the site of zoster (eg, thoracic, lumbar, sacral) that can occur months after resolution of rash points to a long-lasting, virus-induced soluble factor (or factors) that can trigger thrombosis and/or vasculitis. Herein, we investigated the content and contributions of circulating plasma exosomes from HZ and non-HZ patient samples. Compared with non-HZ exosomes, HZ exosomes (1) contained proteins conferring a prothrombotic state to recipient cells and (2) activated platelets leading to the formation of platelet-leukocyte aggregates. Exosomes 3 months after HZ yielded similar results and also triggered cerebrovascular cells to secrete the proinflammatory cytokines, interleukin 6 and 8. These results can potentially change clinical practice through addition of antiplatelet agents for HZ and initiatives to increase HZ vaccine uptake to decrease stroke risk
Convection-Enhanced Drug Delivery: Porous Media Models and Microfluidic Devices
Promising treatments of many brain diseases are often thwarted due to their inability to cross the blood-brain barrier. Convection-enhanced drug delivery (CED) uses direct infusion of drug-containing solutions into tissue to circumvent the blood-brain barrier. The aim of this thesis was to combine models of transport in porous media with microfabricated devices to develop novel methods for controlling the distribution of infused drugs. We used a poroelastic model of the brain to explore the effect of infusion induced dilation on transport. We calculated that during infusions at flow rates greater than one microliter/minute, the effective pore size of the extracellular matrix was doubled by dilation from approximately fifty nanometers to one-hundred nanometers. A computational fluid dynamic model of the rat brain determined the perturbation of the flow field between white and gray matter. We found no further perturbation of the flow field when the ratio of permeabilities between white and gray matter exceeded one-hundred. The results of our models suggest that the material properties of a targeted tissue region dictate the transport of infused solutions. To better control and manipulate drug distribution we developed a novel microfluidic platform for delivering drug-solutions at flow rates relevant for CED. The microfluidic devices consisted of parylene channels with a cross-section area of fifty microns by ten microns on a silicon structure with a cross-sectional area of one-hundred microns by one-hundred microns. These probes were tested in the normal rat brain and demonstrated performance advantages over standard needles, including no channel occlusion and attenuation of backflow. We expanded on the simple single channel device to implement more advanced strategies using multiple channels. A two-channel device was fabricated for infusing an enzyme or mannitol solution prior to infusing polystyrene nanoparticles. By pre-treating the targeted tissue region with enzymes or mannitol we increased the effective pore size of the extracellular matrix which resulted in a doubling of the distribution volume of nanoparticles
Controlled Release Drug Delivery from Hydrogels
The objective of this project is to introduce students to the concepts of diffusion, polymers and polymerization reactions, and enzyme mediated reactions by designing and testing a controlled release drug delivery system. These types of systems are also referred to as localized drug delivery because an implant that releases drugs is placed directly into diseased tissue. Here, we use gelatin as a drug delivery vehicle and food dye as a model drug. Physiological conditions are simulated by placing a cube of the drug delivery system into a test tube with varying ionic and enzymatic conditions. The release of the food dye into the surrounding aqueous environment is measured using a spectrophotomer. This curriculum was developed by Keith Neeves, a graduate student in the School of Chemical and Biomolecular Engineering at Cornell University
Flow chamber and microfluidic approaches for measuring thrombus formation in genetic bleeding disorders
Platelet adhesion and aggregation, coagulation, fibrin formation, and fibrinolysis are regulated by the forces and flows imposed by blood at the site of a vascular injury. Flow chambers designed to observe these events are an indispensable part of doing hemostasis and thrombosis research, especially with human blood. Microfluidic methods have provided the flexibility to design flow chambers with complex geometries and features that more closely mimic the anatomy and physiology of blood vessels. Additionally, microfluidic systems with integrated optics and/or pressure sensors and on-board signal processing could transform what have been primarily research tools into clinical assays. Here, we describe a historical review of how flow-based approaches have informed biophysical mechanisms in genetic bleeding disorders, challenges and potential solutions for developing models of bleeding in vitro, and outstanding issues that need to be addressed prior to their use in clinical settings
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