Antony Thomas

Influence of Red Blood Cells on Nanoparticle Targeted Delivery in Microcirculation

Multifunctional nanomedicine holds considerable promise as the next generation of medicine that allows for targeted therapy with minimal toxicity. Most current studies on Nanoparticle (NP) drug delivery consider a Newtonian fluid with suspending NPs. However, blood is a complex biological fluid composed of deformable cells, proteins, platelets, and plasma. For blood flow in capillaries, arterioles and venules, the particulate nature of the blood needs to be considered in the delivery process. The existence of the cell-free-layer and NP-cell interaction will largely influence both the dispersion and binding rates, thus impact targeted delivery efficacy. In this paper, a particle-cell hybrid model is developed to model NP transport, dispersion, and binding dynamics in blood suspension. The motion and deformation of red blood cells is captured through the Immersed Finite Element Method. The motion and adhesion of individual NPs are tracked through Brownian adhesion dynamics. A mapping algorithm and an interaction potential function are introduced to consider the cell-particle collision. NP dispersion and binding rates are derived from the developed model under various rheology conditions. The influence of red blood cells, vascular flow rate, and particle size on NP distribution and delivery efficacy is characterized. A non-uniform NP distribution profile with higher particle concentration near the vessel wall is observed. Such distribution leads to over 50% higher particle binding rate compared to the case without RBC considered. The tumbling motion of RBCs in the core region of the capillary is found to enhance NP dispersion, with dispersion rate increases as shear rate increases. Results from this study contribute to the fundamental understanding and knowledge on how the particulate nature of blood influences NP delivery, which will provide mechanistic insights on the nanomedicine design for targeted drug delivery applications.

Characterization of nanoparticle delivery in microcirculation using a microfluidic device

This work focuses on the characterization of particle delivery in microcirculation through a microfluidic device. In microvasculature the vessel size is comparable to that of red blood cells (RBCs) and the existence of blood cells largely influences the dispersion and binding distribution of drug loaded particles. The geometry of the microvasculature leads to non-uniform particle distribution and affects the particle binding characteristics. We perform an in vitro study in a microfluidic chip with micro vessel mimicking channels having a rectangular cross section. Various factors that influence particle distribution and delivery such as the vessel geometry, shear rate, blood cells, particle size, particle antibody density are considered in this study. Around 10% higher particle binding density is observed at bifurcation regions of the mimetic microvasculature geometry compared to straight regions. Particle binding density is found to decrease with increased shear rates. RBCs enhance particle binding for both 210 nm and 2 μm particles for shear rates between 200-1600 s(-1) studied. The particle binding density increases about 2-3 times and 6-10 times when flowing in whole blood at 25% RBC concentration compared to the pure particle case, for 210 nm and 2 μm particles respectively. With RBCs, the binding enhancement is more significant for 2 μm particles than that for 210 nm particles, which indicates an enhanced size dependent exclusion of 2 μm particles from the channel centre to the cell free layer (CFL). Increased particle antibody coating density leads to higher particle binding density for both 210 nm and 2 μm particles.

Biomimetic channel modeling local vascular dynamics of pro-inflammatory endothelial changes

Endothelial cells form the inner lining of blood vessels and are exposed to various factors like hemodynamic conditions (shear stress, laminar, and turbulent flow), biochemical signals (cytokines), and communication with other cell types (smooth muscle cells, monocytes, platelets, etc.). Blood vessel functions are regulated by interactions among these factors. The occurrence of a pathological condition would lead to localized upregulation of cell adhesion molecules on the endothelial lining of the blood vessel. This process is promoted by circulating cytokines such as tumor necrosis factor-alpha, which leads to expression of intercellular adhesion molecule-1 (ICAM-1) on the endothelial cell surface among other molecules. ICAM-1 is critical in regulating endothelial cell layer dynamic integrity and cytoskeletal remodeling and also mediates direct cell-cell interactions as part of inflammatory responses and wound healing. In this study, we developed a biomimetic blood vessel model by culturing confluent, flow aligned, endothelial cells in a microfluidic platform, and performed real time in situ characterization of flow mediated localized pro-inflammatory endothelial activation. The model mimics the physiological phenomenon of cytokine activation of endothelium from the tissue side and studies the heterogeneity in localized surface ICAM-1 expression and F-actin arrangement. Fluorescent antibody coated particles were used as imaging probes for identifying endothelial cell surface ICAM-1 expression. The binding properties of particles were evaluated under flow for two different particle sizes and antibody coating densities. This allowed the investigation of spatial resolution and accessibility of ICAM-1 molecules expressed on the endothelial cells, along with their sensitivity in receptor-ligand recognition and binding. This work has developed an in vitro blood vessel model that can integrate various heterogeneous factors to effectively mimic a complex endothelial microenvironment and can be potentially applied for relevant blood vessel mechanobiology studies.

Characterization of vascular permeability using a biomimetic microfluidic blood vessel model

The inflammatory response in endothelial cells (ECs) leads to an increase in vascular permeability through the formation of gaps. However, the dynamic nature of vascular permeability and external factors involved is still elusive. In this work, we use a biomimetic blood vessel (BBV) microfluidic model to measure in real-time the change in permeability of the EC layer under culture in physiologically relevant flow conditions. This platform studies the dynamics and characterizes vascular permeability when the EC layer is triggered with an inflammatory agent using tracer molecules of three different sizes, and the results are compared to a transwell insert study. We also apply an analytical model to compare the permeability data from the different tracer molecules to understand the physiological and bio-transport significance of endothelial permeability based on the molecule of interest. A computational model of the BBV model is also built to understand the factors influencing transport of molecules of different sizes under flow. The endothelial monolayer cultured under flow in the BBV model was treated with thrombin, a serine protease that induces a rapid and reversible increase in endothelium permeability. On analysis of permeability data, it is found that the transport characteristics for fluorescein isothiocyanate (FITC) dye and FITC Dextran 4k Da molecules are similar in both BBV and transwell models, but FITC Dextran 70k Da molecules show increased permeability in the BBV model as convection flow (Peclet number > 1) influences the molecule transport in the BBV model. We also calculated from permeability data the relative increase in intercellular gap area during thrombin treatment for ECs in the BBV and transwell insert models to be between 12% and 15%. This relative increase was found to be within range of what we quantified from F-actin stained EC layer images. The work highlights the importance of incorporating flow in in vitro vascular models, especially in studies involving transport of large size objects such as antibodies, proteins, nano/micro particles, and cells.

Antibody-coated nanoparticles are promising molecular probes for microscopic analysis of cell behavior

Cells are complex systems that respond to local environment and interact with their neighbors through various receptors expressed on the surface. Because of the presence of artifacts associated with microscope images during the fixing process, microscopy assays of fixed and stained cells cannot provide enough information to characterize cellular behaviors and cell–cell interactions [1]. Therefore, development of novel technologies capable of monitoring cellular scale events in real time is of significant importance for providing valuable kinetic and spatial information of cells [2]. Highly sensitive fluorescence microscopy has been widely used for real-time investigation and monitoring of spatiotemporal cellular events such as cell adhesion, cell migration as well as cell–cell, viral–cell, protein–cell and protein–protein interactions [3–5]. For instance, genetically labeled cells with green fluorescent protein (GFP) are being used to visualize cancer cells and metastatic process in live tissue or in the intact animal by whole-body imaging. The development of functional fluorophores and the discovery of GFP-modified versions such as yellow fluorescent protein and cyan fluorescent protein have provided the platform to image several events simultaneously at the single-cell level [1]. One example would be usage of multicolor fluorescence microscopy to study the entry of nanoparticles (NPs) and viruses into living or fixed cells. Dualcolor fluorescent imaging technique is also utilized for subcellular real-time imaging of cancer cell trafficking. By labeling cancer cells with GFP in the nucleus and red fluorescent protein in the cytoplasm, cancer cell trafficking in lymphatic vessels of nude live mice are studied using whole-body imaging. Fluorescent microscopy has further been combined with other microscopy techniques such as atomic force microscopy or electron microscopy to help broaden the understanding of subtle biological interactions and processes [6]. However, the labeling procedures are usually complex and the chemical modification introduced by the fluorescent probe can affect normal cell behavior [7].

Micro-patterned surface for efficient capturing of circulating tumor cells

This work aims to design, fabricate, and characterize a micro-patterned surface that will be integrated into microfluidic devices to enhance particle and rare cell capture efficiency. Capture of ultralow concentration of circulating tumor cells in a blood sample is of vital importance for early diagnostics of cancer diseases. Despite the significant progress achieved in development of cell capture techniques, the enhancement in capture efficiency is still limited and often accompanied with drawbacks such as low throughput, low selectivity, pre-diluting requirement, and cell viability issues. The goal of this work is to design a biomimetic surface that could significantly enhance particle/cell capture efficacy through computational modeling, surface patterning, and microfluidic integration and testing. A PDMS surface with microscale ripples is functionalized with epithelial cell adhesion molecule (EpCAM) antibody to capture prostate cancer PC3 cells. Our microfluidic chip with micropatterns has shown significantly higher cell capture efficiency and selectivity compared to the chips with plane surface or classical herringbone-grooves.

Assemble single stranded DNA and gold nanoparticle complexes onto the surface of RBC

This work aims to assemble single stranded DNA(ssDNA) and gold nanoparticles(AuNPs) into microstructures. The building units of the microstructure first consist of two AuNPs each with one of two sequences of ssDNA attached to their surfaces each with a fluorophore at the free end. A larger piece of hairpin loop ssDNA will then be added with each of its ends complimentary to one of the two ssDNA strands already on the AuNPs. A red blood cell is used as a template for DNA and gold nanoparticles to assemble around. ssDNA-AuNPs binding was confirmed using confocal laser scanning microscope which shows fluorescence in samples after binding.

Characterization of nanoparticle distribution in microcirculation: The influence of blood cells and vascular geometry

This paper focuses on characterization of nanoparticle (NP) distribution in microcirculation. In microcirculation, the vessel size is comparable to the size of red blood cells (RBCs) and the existence of blood cells largely influences the dispersion and binding distribution of NPs. The branching geometry of these vessels leads to non-uniform binding distribution of particles. The influence of various effects such as blood cells, vessel geometry, and shear rates on NP delivery are characterized. The RBCs are found to enhance the binding of NPs at all shear rates while reduces the binding of microparticles at low shear rates. The branching geometry of the capillary vessels leads to higher particle binding density.

A multiphase model for Nanoparticle delivery in microcirculation

In this paper, a particle-cell multiphase model is developed to model Nanoparticle (NP) transport, dispersion, and binding dynamics in blood suspension under the influence of Red blood cells (RBCs). The motion and deformation of RBCs is captured through the Immersed Finite Element Method. The motion and adhesion of individual NPs are tracked through Brownian adhesion dynamics. A mapping algorithm and an interaction potential function are introduced to consider the cell-particle collision. NP dispersion and binding rates are derived from the developed model under various rheology conditions. The influence of RBCs, vascular flow rate, and particle size on NP distribution and delivery efficacy is characterized. A non-uniform NP distribution profile with higher particle concentration near the vessel wall is observed. Such distribution leads to over 50% higher particle binding rate compared to the case without RBC considered. The tumbling motion of RBCs in the core region of the capillary is found to enhance NP dispersion, with dispersion rate increases as shear rate increases. Results from this study contribute to the fundamental understanding and knowledge on how the particulate nature of blood influences NP delivery, which will provide mechanistic insights on the nanomedicine design for targeted drug delivery.

Characterization of Nanoparticle Distribution in Microcirculation Through a Microfluidics Device

Various methods of targeted nano drug delivery have been developed in recent years to reduce side effects, toxicity, and lower drug doses [1]. The use of nanoparticles in drug delivery provides advantages in drug targeting, delivery and release along with serving in diagnosis and therapy [2]. Higher percentage of nanoparticle drug is uptaken by the target cells while larger drug particles are easily cleaned off by the human body. Nanoparticles also have large surface to volume ratio, which aids in attachment of many functional groups and thereby enhances targeting.Copyright