Blood Particulate Analogue Fluids: A Review

Microfluidics has proven to be an extraordinary working platform to mimic and study blood flow phenomena and the dynamics of components of the human microcirculatory system. However, the use of real blood increases the complexity to perform these kinds of in vitro blood experiments due to diverse problems such as coagulation, sample storage, and handling problems. For this reason, interest in the development of fluids with rheological properties similar to those of real blood has grown over the last years. The inclusion of microparticles in blood analogue fluids is essential to reproduce multiphase effects taking place in a microcirculatory system, such as the cell-free layer (CFL) and Fähraeus–Lindqvist effect. In this review, we summarize the progress made in the last twenty years. Size, shape, mechanical properties, and even biological functionalities of microparticles produced/used to mimic red blood cells (RBCs) are critically exposed and analyzed. The methods developed to fabricate these RBC templates are also shown. The dynamic flow/rheology of blood particulate analogue fluids proposed in the literature (with different particle concentrations, in most of the cases, relatively low) is shown and discussed in-depth. Although there have been many advances, the development of a reliable blood particulate analogue fluid, with around 45% by volume of microparticles, continues to be a big challenge.

Manual and Automatic Image Analysis Segmentation Methods for Blood Flow Studies in Microchannels

In blood flow studies, image analysis plays an extremely important role to examine raw data obtained by high-speed video microscopy systems. This work shows different ways to process the images which contain various blood phenomena happening in microfluidic devices and in microcirculation. For this purpose, the current methods used for tracking red blood cells (RBCs) flowing through a glass capillary and techniques to measure the cell-free layer thickness in different kinds of microchannels will be presented. Most of the past blood flow experimental data have been collected and analyzed by means of manual methods, that can be extremely reliable, but they are highly time-consuming, user-intensive, repetitive, and the results can be subjective to user-induced errors. For this reason, it is crucial to develop image analysis methods able to obtain the data automatically. Concerning automatic image analysis methods for individual RBCs tracking and to measure the well known microfluidic phenomena cell-free layer, two developed methods are presented and discussed in order to demonstrate their feasibility to obtain accurate data acquisition in such studies. Additionally, a comparison analysis between manual and automatic methods was performed.

Development of margination of platelet-sized particles in red blood cell suspensions flowing through Y-shaped bifurcating microchannels

BACKGROUND: In the blood flow through microvessels, platelets exhibit enhanced concentrations in the layer free of red blood cells (cell-free layer) adjacent to the vessel wall. The motion of platelets in the cell-free layer plays an essential role in their interaction with the vessel wall, and hence it affects their functions of hemostasis and thrombosis. OBJECTIVE: We aimed to estimate the diffusivity of platelet-sized particles in the transverse direction (the direction of vorticity) across the channel width in the cell-free layer by in vitro experiments for the microchannel flow of red blood cell (RBC) suspensions containing platelet-sized particles. METHODS: Fluorescence microscope observations were performed to measure the transverse distribution of spherical particles immersed in RBC suspensions flowing through a Y-shaped bifurcating microchannel. We examined the development of the particle concentration profiles along the flow direction in the daughter channels, starting from asymmetric distributions with low concentrations on the inner side of the bifurcation at the inlet of the daughter channels. RESULTS: In daughter channels of 40 μm width, reconstruction of particle margination revealed that a symmetric concentration profile was attained in ∼30 mm from the bifurcation, independent of flow rate. CONCLUSIONS: We presented experimental evidence of particle margination developing in a bifurcating flow channel where the diffusivity of 2.9-μm diameter particles was estimated to be ∼40 μm2/s at a shear rate of 1000 s−1 and hematocrit of 0.2.

Emergent cell-free layer asymmetry and biased haematocrit partition in a biomimetic vascular network of successive bifurcations

Blood is a vital soft matter, and its normal circulation in the human body relies on the distribution of red blood cells (RBCs) at successive bifurcations. Understanding how RBCs are...

The effect of hemicylindrical disruptors on the cell free layer thickness in animal blood flows inside microchannels

Blood-side resistance to oxygen transport in extracorporeal membrane blood oxygenators (MBO) depends on fluid mechanics governing the laminar flow in very narrow channels, particularly the hemodynamics controlling the cell free layer (CFL) built-up at solid/blood interfaces. The CFL thickness constitutes a barrier to oxygen transport from the membrane towards the erythrocytes. Interposing hemicylindrical CFL disruptors in animal blood flows inside rectangular microchannels, surrogate systems of MBO mimicking their hemodynamics, proved to be effective in reducing (ca. 20%) such thickness (desirable for MBO to increase oxygen transport rates to the erythrocytes). The blockage ratio (non-dimensional measure of the disruptor penetration into the flow) increase is also effective in reducing CFL thickness (ca. 10–20%), but at the cost of risking clot formation (undesirable for MBO) for disruptors with penetration lengths larger than their radius, due to large residence times of erythrocytes inside a low-velocity CFL formed at the disruptor/wall edge.

Stiff Erythrocyte Subpopulations Biomechanically Induce Endothelial Inflammation in Sickle Cell Disease

Background: Originally described as a monogenic hemoglobin disorder resulting in increased red blood cell (RBC) stiffness leading to vaso-occlusion, sickle cell disease (SCD) is now known to be a vasculopathic disease with some semblance to cardiovascular disease in which the endothelium is inflamed. While adhesive RBC-endothelial interactions, inflammatory cytokines, and hemolysis all contribute to SCD vasculopathy, whether the increased stiffness of sickle RBCs directly contributes to endothelial inflammation is unknown. Endothelial cells are now known to mechanotransduce shear forces into biological signals. Pathological alteration of such forces leads to proinflammatory endothelial cell signaling including upregulation of VCAM-1 and E-selectin, which contribute to atherosclerotic plaques leading to myocardial infarction and stroke (Abe, ATVB, 2014). In addition, under normal homeostatic conditions, RBCs do not come into contact with the endothelium due to a cell-free layer created by the Fåhræus-Lindqvist effect. Studies including our own have shown in silico that increasing RBC stiffness diminishes or eliminates the cell-free layer, allowing stiff RBCs to contact the vessel wall (Kumar, Phys Rev E, 2011). This is particularly pertinent in SCD, as all patients have a small population (1-10%) of sickle RBCs that are permanently stiff and misshapen. We therefore hypothesize that purely physical interactions - akin to "scratches" or collisions - between endothelial cells and stiff SCD RBCs breaking through the cell-free layer are sufficient to cause endothelial inflammation in the absence of adhesion or vaso-occlusion (Fig. 1A). Methods: We performed computational direct numerical simulations using the boundary integral method for a binary suspension of flexible biconcave discs and stiff curved prolate spheres modeling healthy RBCs and ISCs, respectively. Experimentally, we leveraged our microfluidic microvasculature models of human umbilical vein endothelial cells cultured throughout each microchannel (Fig. 2). RBCs from SCD patients were "spiked" into normal RBC suspensions to comprise 5 and 10% of the overall population (a representation of ISCs in vivo), suspended in media to 25% hematocrit mimicking conditions seen in SCD patients, and perfused into the microfluidics for 4 hours. Samples of 100% normal RBCs or SCD RBCs were run in parallel. To isolate the stiffness effects of sickle RBCs without confounding hemolytic and adhesive effects, parallel experiments were conducted using nystatin-treated normal RBCs to create artificially stiffened RBC subpopulations, defined by elevated mean corpuscular hemoglobin concentrations (MCHCs), at the same proportion of the overall RBC population (0, 5, 10 and 100%). The endothelialized models were then fixed, permeabilized, and immunostained with antibodies against VCAM-1 and E-selectin. Mean fluorescence intensity was measured to quantify endothelial inflammation. Results: In silico, we observed that ISCs strongly marginate towards the vessel walls due to their stiffness and "pointy" shape, and heterogeneous suspensions with small fractions of stiff, pointy cells (5 and 10%) caused the highest degree of margination (Fig. 1B). Experimentally, endothelium exposed to 5, 10, and 100% SCD RBCs exhibited increased VCAM-1 and E-selectin expression over normal RBCs, and the degree of expression increased with higher percentages of SCD RBCs. While endothelial cells exposed to nystatin-stiffened RBCs also showed increased VCAM-1 and E-selectin expression, those exposed to a lower percentage of stiff cells (5 and 10%) exhibited higher expression than the homogenously stiff (100%) condition (Fig. 3), which is consistent with our computer simulations. Conclusions: Here we demonstrate that purely non-adhesive, physical interactions between endothelial cells and SCD RBCs are sufficient to cause endothelial inflammation. Furthermore, heterogeneous RBC populations, comprised of a small minority of stiff cells, cause more inflammation than uniformly stiff RBCs. Studies elucidating the underlying mechanisms, using different endothelial cell types, and analyzing the effect of vessel curvature are ongoing. Our results introduce a new paradigm for understanding SCD pathophysiology and may help explain how chronic diffuse vasculopathy develops, which could lead to more biophysically-based therapeutic strategies. Disclosures Carden: GBT: Honoraria; NIH: Research Funding. Mannino:Sanguina, LLC: Employment, Equity Ownership. Lam:Sanguina, LLC: Equity Ownership.

Experimental and Numerical Study of Blood Flow in μ-vessels: Influence of the Fahraeus–Lindqvist Effect

The study of hemodynamics is particularly important in medicine and biomedical engineering as it is crucial for the design of new implantable devices and for understanding the mechanism of various diseases related to blood flow. In this study, we experimentally identify the cell free layer (CFL) width, which is the result of the Fahraeus–Lindqvist effect, as well as the axial velocity distribution of blood flow in microvessels. The CFL extent was determined using microscopic photography, while the blood velocity was measured by micro-particle image velocimetry (μ-PIV). Based on the experimental results, we formulated a correlation for the prediction of the CFL width in small caliber (D < 300 μm) vessels as a function of a modified Reynolds number (Re∞) and the hematocrit (Hct). This correlation along with the lateral distribution of blood viscosity were used as input to a “two-regions” computational model. The reliability of the code was checked by comparing the experimentally obtained axial velocity profiles with those calculated by the computational fluid dynamics (CFD) simulations. We propose a methodology for calculating the friction loses during blood flow in μ-vessels, where the Fahraeus–Lindqvist effect plays a prominent role, and show that the pressure drop may be overestimated by 80% to 150% if the CFL is neglected.

Effect of Cytoplasmic Viscosity on Red Blood Cell Migration in Small Arteriole-level Confinements

The dynamics of red blood cells in small arterioles are important as these dynamics affect many physiological processes such as hemostasis and thrombosis. However, studying red blood cell flows via computer simulations is challenging due to the complex shapes and the non-trivial viscosity contrast of a red blood cell. To date, little progress has been made studying small arteriole flows (20-40μm) with a hematocrit (red blood cell volume fraction) of 10-20% and a physiological viscosity contrast. In this work, we present the results of large-scale simulations that show how the channel size, viscosity contrast of the red blood cells, and hematocrit affect cell distributions and the cell-free layer in these systems. We utilize a massively-parallel immersed boundary code coupled to a finite volume solver to capture the particle resolved physics. We show that channel size qualitatively changes how the cells distribute in the channel. Our results also indicate that at a hematocrit of 10% that the viscosity contrast is not negligible when calculating the cell free layer thickness. We explain this result by comparing lift and collision trajectories of cells at different viscosity contrasts.