Effect of Material Type and Particle Density On the Margination of Vascular-Targeted Carriers

Effect of Material Type and Particle Density on the Margination of Vascular-Targeted Carriers

Alex Thompson¹ and Omolola Eniola-Adefeso¹

(1) Chemical Engineering, University of Michigan, Ann Arbor, MI

Introduction:   The development of vascular-targeted carriers (VTC) for the localized delivery of therapeutics could greatly improve the treatment of many human diseases.  The ability to deliver a high fraction of an injected dose to a targeted diseased area is the paramount goal of vascular targeting.  In order for VTCs to be effective, they must be able to marginate (localize and adhere) to diseased endothelium from blood flow. Recent research has shown that particle size and shape affect particle margination in the presence of red blood cells.^1,2,3 Neutrally buoyant microparticles (relative to blood) with equivalent spherical diameter (ESD)  ³ 2 µm undergo a forced localization to the red blood cell free layer (RBC-FL); analogous to the mechanism by which red blood cells force leukocytes and platelets to the RBC-FL, allowing them to effectively interact with the endothelium.  Smaller, neutrally buoyant spherical particles (100-1000 nm ESD) do not benefit from this effect, therefore making them suboptimal for use as VTCs, particularly in treating diseases that occur in M/LBV, such as atherosclerosis. VTCs can be made from a variety of materials which have densities different from that of blood. The purpose of this study is to examine how the size and density of potential VTCs prescribe their ability to localize the RBC-FL in vitro in human blood flow under physiological conditions.

Materials and Methods:  Fluorescent spherical particles of different material with different densities were conjugated with targeting molecules (sialyl Lewis a, sLe^a) which bind to selectins at a fixed surface density.  Targeted particles were tested for their margination efficiency from blood flow in vitro using parallel plate flow chamber (PPFC) assays.  The particles explored ranged in diameter from 100-1000 nm.

vascular-targeted spheres at a fixed concentration were mixed with blood from human donors and allowed to flow over a layer of IL-1β activated human umbilical vein endothelial cells (HUVEC) lining one wall of a parallel plate flow chamber. Adhesion of VTCs from laminar, steady blood flow from a range of wall shear rates (200-1000 s^-1) was investigated.  Channel orientation was varied in four different configurations in relation to gravity to examine how particles of different densities marginate from blood flow.  Particle adhesion in recirculating blood flow, in which a sudden expansion in the PPFC causes a region of recirculating flow to occur in the channel, was also examined to see if a disturbed flow profile helps particles of different densities from blood separate from the RBC core more efficiently.  The number of adherent particles under various flow conditions was imaged/quantified using brightfield and fluorescence microscopy. Also, the localization to the RBC-FL during blood flow was visualized using confocal microscopy.

Results and Discussion:   Results from in vitro flow assays show that, as expected, the adhesion of neutrally buoyant polystyrene (PS) microspheres was not affected by chamber orientation.  Silica (Si) microspheres with 1 µm diameter display roughly double the adhesion of equivalent polystyrene (PS) spheres when the chamber was oriented with the HUVEC monolayer at the bottom; however, the same Si microspheres adhere at roughly the same level as PS microspheres when the chamber is inverted so that the monolayer is at the top of the chamber. When the target site is in the downward direction, gravity favors the more dense Si particle in terms of adhesion; however when the target site is upward, perhaps the presence of the RBC core below allows similar levels of Si microparticles to remain in the RBC-FL to interact with the endothelium. In recirculation flow, our results show that at lower shear rates (200 s^-1) Si microparticles adhere at higher levels than PS microparticles whether the chamber is oriented with the monolayer at the top or the bottom.  It appears that the flow normal to the monolayer caused by the recirculation region helps Si redistribute to the RBC-FL in this disturbed flow, resulting in an improved adhesion in this area.  This finding could have implication in targeting atherosclerosis, which is known to occur in areas of the vasculature where this type of disturbed flow is found.  Ongoing work is investigating the margination smaller (20-200 nm) and more dense (FeO, Au, etc) particles with the goal of identifying particle characteristics which would make them suitable for targeting specific areas of the vasculature. 

Conclusions:   Our study shows that particle density can affect the margination of vascular-targeted carriers, which can be particularly useful in designing carriers which target diseases, such as atherosclerosis, which occur in areas of the vasculature that exhibit unique hemodynamics.