Fluid Dynamic Environment Research Articles

COMPUTATIONAL FLUID DYNAMICS FOR TISSUE ENGINEERING APPLICATIONS

Hydrodynamic cellular environment plays an important role in translating engineered tissue constructs into clinically useful grafts. However, the cellular fluid dynamic environment inside bioreactor systems is highly complex and it is normally impractical to experimentally characterize the local flow patterns at the cellular scale. Computational fluid dynamics (CFD) has been recognized as an invaluable and reliable alternative to investigate the complex relationship between hydrodynamic environments and the regeneration of engineered tissues at both the macroscopic and microscopic scales. This review describes the applications of CFD simulations to probe the hydrodynamic environment parameters (e.g., flow rate, shear stress, etc.) and the corresponding experimental validations. We highlight the use of CFD to optimize bioreactor design and scaffold architectures for improved ex-vivo hydrodynamic environments. It is envisioned that CFD could be used to customize specific hydrodynamic cellular environments to meet the unique requirements of different cell types in combination with advanced manufacturing techniques and finally facilitate the maturation of tissue-engineered constructs.

A computational and experimental study inside microfluidic systems: the role of shear stress and flow recirculation in cell docking

In this paper, microfluidic devices containing microwells that enabled cell docking were investigated. We theoretically assessed the effect of geometry on recirculation areas and wall shear stress patterns within microwells and studied the relationship between the computational predictions and experimental cell docking. We used microchannels with 150 microm diameter microwells that had either 20 or 80 microm thickness. Flow within 80 microm deep microwells was subject to extensive recirculation areas and low shear stresses (<0.5 mPa) near the well base; whilst these were only presented within a 10 microm peripheral ring in 20 microm thick microwells. We also experimentally demonstrated that cell docking was significantly higher (p < 0.01) in 80 microm thick microwells as compared to 20 microm thick microwells. Finally, a computational tool which correlated physical and geometrical parameters of microwells with their fluid dynamic environment was developed and was also experimentally confirmed.

Cell Distribution in a Scaffold with Random Architectures under the Influence of Fluid Dynamics

Fluid dynamic environment and scaffold architectures have an important influence on cell growth and distribution inside the scaffold. A porous cylindrical scaffold with a central channel is seeded with the sheep mesenchymal stem cells (MSCs) in this study. Then the cell seeded scaffold is continuously perfused with alpha-MEM medium by a peristaltic pump for 7, 14, and 28 days. Histological study shows that the cell proliferation rates are different throughout the whole scaffolds. The different cell coverage is shown in various positions of the scaffold. A computational fluid dynamics (CFD) modeling is used to simulate the flow conditions within perfused cell-seeded scaffolds to give insight into the mechanisms of these cell growth phenomena. Relating the simulation results to perfusion experiments, the even fluid velocity (approximately 0.26-0.64 mm/s) and shear stress (approximately 0.0029-0.027 Pa) are found to correspond to increased cell proliferation within the cell-scaffold constructs. This method exhibits novel capabilities to compare results obtained for different perfusion rates or different scaffold microarchitectures and may allow specific fluid velocities and shear stresses to be determined that optimize the perfusion flow rate, porous scaffold architecture, and distribution of in vitro tissue growth.

Microcirculation within grooved substrates regulates cell positioning and cell docking inside microfluidic channels

Immobilization of cells inside microfluidic devices is a promising approach for enabling studies related to drug screening and cell biology. Despite extensive studies in using grooved substrates for immobilizing cells inside channels, a systematic study of the effects of various parameters that influence cell docking and retention within grooved substrates has not been performed. We demonstrate using computational simulations that the fluid dynamic environment within microgrooves significantly varies with groove width, generating microcirculation areas in smaller microgrooves. Wall shear stress simulation predicted that shear stresses were in the opposite direction in smaller grooves (25 and 50 microm wide) in comparison to those in wider grooves (75 and 100 microm wide). To validate the simulations, cells were seeded within microfluidic devices, where microgrooves of different widths were aligned perpendicularly to the direction of the flow. Experimental results showed that, as predicted, the inversion of the local direction of shear stress within the smaller grooves resulted in alignment of cells on two opposite sides of the grooves under the same flow conditions. Also, the amplitude of shear stress within microgrooved channels significantly influenced cell retainment in the channels. Therefore, our studies suggest that microscale shear stresses greatly influence cellular docking, immobilization, and retention in fluidic systems and should be considered for the design of cell-based microdevices.

Potential Effect of Geometry on Wall Shear Stress Distribution Across Scaffold Surfaces

Bioreactors are used as cell culture systems for growth and maintenance of tissue-engineered scaffolds that serve as three-dimensional (3D) templates for initial cell attachment and subsequent tissue formation. The bioreactors' fluid dynamic environment is known to play a crucial role in the synthesis of cellular components via flow-mediated mechanical stimuli. Computational fluid dynamics (CFD) models in the slow turning lateral vessel (STLV Synthecon, Inc.) were simulated under Couette flow conditions. Systematic research of the wall shear stress (WSS) effects on the scaffold's geometry has been limited. Therefore, direct qualitative and quantitative correlations for WSS values were performed by analyzing and comparing WSS value distributions of two scaffold shapes. Under experimental flow conditions, the disc and prolate spheroid shapes exhibited dissimilar WSS distribution. Nonetheless, when compared to the disc models, the high pressure stagnation region of the spheroid was reduced between 60% and 95%. In the spheroid shape, approximately 40% increase in the shear stress surface exposure to flow ranged from 2 to 3 dyn/cm(2). These values suggest that WSSs are likely affected by scaffold shape and vary little with location within the Synthecon STLV. The proposed simulation studies evidenced the CFD model's flexibility to characterize and quantify forces affecting tissue-engineered scaffold design.

The growth of stem cells within β-TCP scaffolds in a fluid-dynamic environment

A three-dimensional dynamic perfusion system was developed to provide mass transport and nutrient supply to permit the cell proliferation during the long-term culture inside a β-tricalcium phosphate (β-TCP) scaffold. Also the flow field throughout the scaffold was studied. The porous cylindrical scaffold with a central channel was seeded with the sheep mesenchymal stem cells (MSCs). Then the cell-seeded scaffolds were continuously perfused with the complete α-MEM medium by a peristaltic pump for 7, 14 and 28 days, respectively. Histological study showed that the cell proliferation rates were different throughout the whole scaffolds and the different cell coverage was shown in different positions of the scaffold. Unoccupied spaces were found in many macropores. A computational fluid dynamics (CFD) modeling was used to simulate the flow conditions within perfused cell-seeded scaffolds to give an insight into the mechanisms of these cell growth phenomena. Relating the simulation results to perfusion experiments, the even fluid velocity (approximately 0.52 mm/s) and shear stress (approximately 0.0055 Pa) were found to correspond to increased cell proliferation within the cell–scaffold constructs. Flow speeds were between 0.25 and 0.75 mm/s and shear stresses were between 0.003 and 0.008 Pa in approximately 75% of the regions. This method exhibits novel capabilities to compare the results obtained for different perfusion rates or different scaffold microarchitectures. It may allow specific fluid velocities and shear stresses to be determined to optimize the perfusion flow rate, porous scaffold architecture and distribution of in vitro tissue growth.

Flow modelling within a scaffold under the influence of uni-axial and bi-axial bioreactor rotation

The problem of donor scarcity has led to the recent development of tissue engineering technologies, which aim to create implantable tissue equivalents for clinical transplantation. These replacement tissues are being realised through the use of biodegradable polymer scaffolds; temporary/permanent substrates, which facilitate cell attachment, proliferation, retention and differentiated tissue function. To optimise gas transfer and nutrient delivery, as well as to mimic the fluid dynamic environment present within the body, a dynamic system might be chosen. Experiments have shown that dynamic systems enhance tissue growth, with the aid of scaffolds, as compared to static culture systems. Very often, tissue growth within scaffolds is only seen to occur at the periphery. The present study utilises the Computational Fluid Dynamics package FLUENT, to provide a better understanding of the flow phenomena in scaffolds, within our novel bioreactor system. The uni-axial and bi-axial rotational schemes are studied and compared, based on a vessel rotating speed of 35 rpm. The wall shear stresses within and without the constructs are also studied. Findings show that bi-axial rotation of the vessel results in manifold increases of fluid velocity within the constructs, relative to uni-axial rotation about the X- and Z-axes, respectively.

Oxygen transport and cell viability in an annular flow bioreactor: Comparison of laminar Couette and Taylor‐vortex flow regimes

Rotating wall vessel bioreactors have been proposed as a means of controlling the fluid dynamic environment during long-term culture of mammalian cells and engineered tissues. In this study, we show how the delivery of oxygen to cells in an annular flow bioreactor is enhanced by the forced convective transport afforded by Taylor vortex flows. A fiberoptic oxygen probe with negligible lag time was used to measure the dissolved oxygen concentration in real time and under carefully controlled aeration conditions. From these data, the overall mass transfer coefficients were calculated and mass transport correlations determined under laminar Couette flow conditions and discrete Taylor vortex flow regimes, including laminar, wavy, and turbulent flows. While oxygen transport in Taylor vortex flows was significantly greater, and the available oxygen exceeded that consumed by murine fibroblasts in free suspension, the proportion of cells that remained viable decreased with increasing Reynolds number (101.8 < Rei < 1018), which we attribute to the action of fluid shear stresses on the cells as opposed to any limitation in mass transport. Nevertheless, the results of this study suggest that laminar Taylor-vortex flow regimes provide an effective means of maintaining the levels of oxygen transport required for long-term cell culture.

Analysis of flow instabilities in convex and concave floating zones heated by an equatorial ring under microgravity conditions

This analysis deals with advances in models concerning the floating zone technique as well as with novel results on the relative importance of various parameters in the crystal-growth process. The attention is focused in particular on microgravity fluid-dynamic aspects and on the effect of the volume of the liquid melt since the cylindrical configuration is expected to be only a very special case under microgravity conditions. The instability of Marangoni flow is investigated by direct three-dimensional and time-dependent simulation of the problem and parallel computations. Body-fitted curvilinear co-ordinates are adopted to handle the non-cylindrical shape. A novel realistic distribution is considered to model the surface heat flux generated by a ring heater positioned around the equatorial plane at a fixed distance from the axis of the liquid (full) zone. The fluid-dynamic environment that occurs inside the melt is very sensitive to the geometrical aspect ratio A F (length/diameter) of the floating zone and to the volume factor S (ratio of the volume of the liquid zone and the volume of the corresponding cylindrical configuration: convex S>1, concave S<1). The role played by the geometrical constraints and degrees of freedom of the Marangoni toroidal rolls in determining the azimuthal structure and the stability of the flow field is discussed.

Rho Mediates the Shear-Enhancement of Endothelial Cell Migration and Traction Force Generation

The migration of vascular endothelial cells in vivo occurs in a fluid dynamic environment due to blood flow, but the role of hemodynamic forces in cell migration is not yet completely understood. Here we investigated the effect of shear stress, the frictional drag of blood flowing over the cell surface, on the migration speed of individual endothelial cells on fibronectin-coated surfaces, as well as the biochemical and biophysical bases underlying this shear effect. Under static conditions, cell migration speed had a bell-shaped relationship with fibronectin concentration. Shear stress significantly increased the migration speed at all fibronectin concentrations tested and shifted the bell-shaped curve upwards. Shear stress also induced the activation of Rho GTPase and increased the traction force exerted by endothelial cells on the underlying substrate, both at the leading edge and the rear, suggesting that shear stress enhances both the frontal forward-pulling force and tail retraction. The inhibition of a Rho-associated kinase, p160ROCK, decreased the traction force and migration speed under both static and shear conditions and eliminated the shear-enhancement of migration speed. Our results indicate that shear stress enhances the migration speed of endothelial cells by modulating the biophysical force of tractions through the biochemical pathway of Rho-p160ROCK.

Correlation of Intimal Hyperplasia Development and Shear Stress Distribution at the Distal End-side-anastomosis, in vitro Study Using Particle Image Velocimetry

Low shear areas at the distal anastomosis of peripheral bypasses are thought to promote neointimal hyperplasia. In this study we evaluated the fluid dynamic environment at the distal anastomosis of peripheral bypasses by means of a new method for in vitro flow visualization and quantitative velocity field measurement.A silastic model of a distal end-side anastomosis was attached to a mock circulation loop driven by an artificial heart. High resolution velocity fields were measured by means of particle image velocimetry (PIV). The velocity vector data were used to calculate vorticity ω, strain rates ex, shear rates h and shear stresses τ. Two separations and a stagnation zone were identified by means of flow visualization. Measured velocities inside the three zones were significantly lower than in the high velocity mainstream. Calculated shear rates and shear stresses inside the zones were significantly lower than human wall shear rates. At the transition between the effective mainstream and the boundary layers high vorticity and compressive strain fields existed, indicating the presence of high shear forces. The locations of these areas corresponded to the well known zones of intimal hyperplasia.The high resolution shear stress analysis supports the low shear theory of intimal hyperplasia development. A wall diversion angle greater than 6° leads to flow separation and presumed IH promotion until high shear transition areas are reached.

Mitral mechanical heart valves: in vitro studies of their closure, vortex and microbubble formation with possible medical implications.

The goal of the present work was to create the closest possible in vitro fluid dynamic environment in which prosthetic mitral valves in the patients' hearts function, in order to demonstrate whether microbubbles are generated, and if yes, under what conditions and at which stage of the cardiac cycle. Microbubbles were observed in the blood of patients with mitral mechanical heart valves (MHV) by means of echocardiography. The phenomenon, often referred to as high-intensity transient signals (HITS), appears as bright, intense, high-velocity and persistent echoes detected by Doppler echocardiography at the instant of valve closure. The question is no longer whether microbubbles are being formed in patients with MHV. as an inherent aspect of their design, but rather how they evolve and when. The answer to this question was the objective of the present paper. Hemodynamic conditions in which microbubbles were observed in patients with mitral MHV were simulated in our laboratory. We were able to describe the bubble formation process, as one consisting of nucleation and microbubble growth. While mild growth of nuclei is governed by diffusion, extensive growth of microbubbles is controlled by pressure drop during deceleration of the leaflets on the housing on the atrial side of the mitral MHV. The present study has shown that bubbles form in a fluid at the instant of closure of mechanical valves. The formation of vortices after valve closure, although clinically not yet observed, was also demonstrated in the present in vitro studies. We believe that impact of such vortices on the endothelial layer of the left atrial wall may have clinical significance. These two phenomena were not observed in bioprosthetic valves. As demonstrated, there exist two distinct phenomena characteristic of mechanical heart valves, which take place during valve closure, namely, that of vortex formation and that of microbubble growth. Both phenomena may have far reaching clinical implications.

The growth and the fluid dynamics of protein crystals and soft organic tissues: models and simulations, similarities and differences

The fluid-dynamic environment within typical growth reactors as well as the interaction of such flow with the intrinsic kinetics of the growth process are investigated in the frame of the new fields of protein crystal and tissue engineering. The paper uses available data to introduce a set of novel growth models. The surface conditions are coupled to the exchange mass flux at the specimen/culture-medium interface and lead to the introduction of a group of differential equations for the nutrient concentration around the sample and for the evolution of the construct mass displacement. These models take into account the sensitivity of the construct/liquid interface to the level of supersaturation in the case of macromolecular crystal growth and to the “direct” effect of the fluid-dynamic shear stress in the case of biological tissue growth. They then are used to show how the proposed surface kinetic laws can predict (through sophisticated numerical simulations) many of the known characteristics of protein crystals and biological tissues produced using well-known and widely used reactors. This procedure provides validation of the models and associated numerical method and at the same time gives insights into the mechanisms of the phenomena. The onset of morphological instabilities is discussed and investigated in detail. The interplay between the increasing size of the sample and the structure of the convective field established inside the reactor is analysed. It is shown that this interaction is essential in determining the time evolution of the specimen shape. Analogies about growing macromolecular crystals and growing biological tissues are pointed out in terms of behaviours and cause-and-effect relationships. These aspects lead to a common source (in terms of original mathematical models, ideas and results) made available for the scientific community under the optimistic idea that the contacts established between the “two fields of engineering” will develop into an ongoing, mutually beneficial dialogue.

Ligand coated nanosphere adhesion to E- and P-selectin under static and flow conditions.

The heterogeneous distribution of endothelial cell adhesion molecules (ECAMs) on the lumenal surface of vascular endothelium provides an opportunity to deliver drugs to select tissues. The targeting could be achieved by using carriers whose outer surface has a ligand for a selectively expressed ECAM. The carriers would interact with the endothelium in a fluid dynamic environment and in many of these schemes nanoparticles would be used. It is unclear what role various parameters (e.g., ligand-ECAM chemistry, fluid shear) will have on the adhesion of the nanoparticles to the endothelium. To facilitate studies in this area, we have developed a prototypical in vitro model that allows investigation of nanoparticle adhesion. We coated polystyrene nanospheres with a humanized mAb (HuEP5C7.g2) that recognizes the ECAMs E- and P-selectin. Adhesion assays revealed that HuEP5C7.g2 nanospheres exhibit augmented, specific adhesion to selectin presenting cellular monolayers and that the adhesion can be affected by the fluid shear. These results; (i) strongly suggest that HuEP5C7.g2 could be used to target nanoparticles to selectin presenting endothelium; (ii) demonstrate that fluid shear can affect nanoparticle adhesion; and (iii) define a system which can be used to study the effects of various system parameters on nanoparticle adhesion.

The fluid dynamic and shear environment in the NASA/JSC rotating-wall perfused-vessel bioreactor.

The rotating-wall perfused-vessel (RWPV) bioreactor, used for both microgravity and Earth-based cell science experiments, is characterized in terms of the fluid dynamic and fluid shear stress environment. A numerical model of the flow field is developed and verified with laser Doppler velocimeter measurements. The effects of changes in operating conditions, including rotation rates and fluid perfusion rates, are investigated with the numerical model. The operating conditions typically used for ground-based experiments (equal rotation of the inner and outer cylinders) leads to flow patterns with relatively poor mass distribution characteristics. Approximately 50% of the inlet-perfused fluid bypasses the bulk of the fluid volume and flows to the perfusion exit. For operating conditions typical in microgravity, small differential rotation rates between the inner and outer cylinders lead to greatly improved flow distribution patterns and very low fluid shear stress levels over a large percentage of the fluid volume. Differences in flow patterns for the different operating conditions are explored. Large differences in the hydrodynamic environments for operating conditions typical of true microgravity and ground-based "microgravity simulations" are demonstrated.

Cell-cell adhesive interactions in an in vitro flow chamber.

A critical step in the recruitment of leukocytes to a site of inflammation is leukocyte adhesion to the vascular endothelium in the fluid dynamic environment of the microcirculation. This process involves a cascade of adhesive events, including initial attachment, rolling, spreading, and ultimately transendothelial migration (reviewed in ,). To elucidate the biochemistry and biophysics that govern these processes, in vitro flow chamber assays have been developed. These assays allow the study of leukocyte adhesion under well-defined fluid flow conditions that mimic the in vivo fluid dynamic environment. The parallel plate flow chamber is the design most frequently used for this purpose. Such a device was used in the late 1980s to study human neutrophil adhesion to cultured human umbilical vein endothelial monolayers (). This group (,) has recently published a report that details the chamber design and operation. In general, this method involves perfusing a suspension of purified leukocytes over a coverslip containing a monolayer of cells or a region of purified adhesion molecules. Adhesive events are monitored in live time and recorded by videomicroscopy for later analysis. We present here the methodology of using the flow chamber currently used by our group to study leukocyte and tumor cell adhesion to vascular endothelium or purified adhesion molecules under defined laminar flow conditions.

Odor Plumes and Animal Navigation in Turbulent Water Flow: A Field Study.

Turbulence causes chemical stimuli to be highly variable in time and space; hence the study of animal orientation in odor plumes presents a formidable challenge. Through combined chemical and physical measurements, we characterized the transport of attractant released by clam prey in a turbulent aquatic environment. Concurrently, we quantified the locomotory responses of predatory crabs successfully searching for sources of clam attractant. Our results demonstrate that both rheotaxis and chemotaxis are necessary for successful orientation. Perception of chemical cues causes crabs to move in the upstream direction, but feedback from attractant distributions directly regulates movement across-stream in the plume. Orientation mechanisms used by crabs difler from those employed by flying insects, the only other system in which navigation relative to odor plumes has been coupled with fluid dynamics. Insects respond to odors by moving upstream, but they do not use chemical distributions to determine across-stream direction, whereas crabs do. Turbulent eddy diffusivities in crab habitats are 100 to 1000 times lower than those of terrestrial grasslands and forests occupied by insects. Insects must respond to plumes consisting of highly dispersed, tiny filaments or parcels of odor. Crabs rely more heavily on spatial aspects of chemical stimulus distributions because their fluid dynamic environment creates a more stable plume structure, thus permitting chemotaxis.