Evaluating Nanodisks and Nanorods As Carriers for Targeting Diseased Endothelium in Physiological Blood Flows

Introduction:   The development of vascular-targeted carriers (VTC) for the delivery of therapeutics could greatly improve the treatment of many human diseases. Spherical nanoparticles are commonly used as potential drug carriers due to their ability to easily navigate microvasculature and ease of fabrication/drug loading. However, recent literature has shown that spherical nanoparticles do not efficiently marginate and adhere to the vascular wall in model blood vessels compared to larger micron sized particles.^1,2 Therefore spherical nanoparticles may not be effective as VTCs for drug delivery and diagnosis, particularly in targeting diseases which affect medium/large blood vessels (M/LBV), such as atherosclerosis. Recently, particle shape has received attention as a parameter that can be used to improve the performance of VTCs. The purpose of this study is to examine the hemodynamics of rod and disk-shaped particles of different aspect ratios relative to spheres of equal volume both in vitro in human blood flow under physiological conditions.

Materials and Methods:  Polystyrene rod and disk-shaped particles were fabricated via a polymer film stretch method previously described in literature.³ Rods, disks, and spherical particles were conjugated with targeting molecules (sialyl Lewis a, sLe^a) which bind to selectins.  The differently shaped particles were tested for their margination efficiency from blood flow both in vitro and in vivo.  The particles explored ranged in diameter from 0.5 – 2 μm (equivalent spherical diameter; ESD, for rods and disks).

Vascular-targeted spheres, rods or disks at a fixed concentration were mixed with RBC and saline flow buffer, and allowed to flow over a layer of IL-1β activated human umbilical vein endothelial cells (HUVEC) using a parallel plate flow chamber. Adhesion of VTCs from steady blood flow from relatively low to relatively high shear rates (200-1000 s^-1) was investigated. Since disturbed blood flow profiles are typical in areas of vasculature where atherosclerosis tends to occur, particle adhesion pulsatile and recirculatory blood flow profiles were also examined.  The adhesion of rods, disks and spheres to the endothelium from blood flow was imaged/quantified using brightfield and fluorescence microscopy. Further, the localization of these particles to the red blood cell free layer (RBC-FL) while in flow was investigated using confocal microscopy.

Results and Discussion:  With in vitro assays we find that rod-shaped microparticles at a high aspect ratio have a higher binding efficiency to the HUVEC monolayer from human blood flow than spheres of equivalent volume and targeting ligand site density. Overall, adhesion was greatest for microparticles with ESD=2 μm compared to those with smaller volumes.  The improved adhesion pattern for micron-sized rods was not due to better localization of rods to the RBC-FL, as the confocal data showed equivalent levels of microrods/spheres in the RBC-FL.  Adhesion of 500 nm ESD spheres and rods was minimal; likely due to poor localization to the RBC-FL.  While rods with 500 nm ESD showed no improved adhesion compared to equivalent spheres, preliminary data suggests that equivalent disk shaped particles display both better localization to the RBC-FL and improved adhesion to the endothelium. 

Conclusions:   Our study shows that shape can be a very useful and tunable parameter in the design of vascular-targeted carriers for imaging and treatment of atherosclerosis and other diseases of M/LBVs.