Roger Tran-Son-Tay

Computational modeling of cell adhesion and movement using a continuum-kinetics approach

Adhesion of leukocytes to substrate involves the coupling of disparate length and timescales between molecular mechanics and macroscopic transport, and existing models of cell adhesion do not use full cellular information. To address these challenges, a multiscale computational approach for studying the adhesion of a cell on a substrate is developed and assessed. The cellular level model consists of a continuum representation of the field equations and a moving boundary tracking capability to allow the cell to change its shape continuously. At the receptor-ligand level, a bond molecule is mechanically represented by a spring. Communication between the macro/micro- and nanoscale models is facilitated interactively during the computation. The computational model is assessed using an adherent cell, rolling and deforming along the vessel wall under imposed shear flows. Using this approach, we first confirm existing numerical and experimental results. In this study, the intracellular viscosity and interfacial tension are found to directly affect the rolling of a cell. Our results also show that the presence of a nucleus increases the bond lifetime, and decreases the cell rolling velocity. Furthermore, it is found that a cell with a larger diameter rolls faster, and decreases the bond lifetime. This study shows that cell rheological properties have significant effects on the adhesion process contrary to what has been hypothesized in most literature.

Determination of red blood cell membrane viscosity from rheoscopic observations of tank-treading motion

Measurements of the dimensions and membrane rotational frequency of individual erythrocytes steadily tank-treading in a rheoscope are used to deduce the surface shear viscosity of the membrane. The method is based on an integral energy principle which says that the power supplied to the tank-treading cell by the suspending fluid is equal to the rate at which energy is dissipated by viscous action in the membrane and cytoplasm. The integrals involved are formulated with the aid of an idealized mathematical model of the tank-treading red blood cell (RBC) (Keller and Skalak, 1982, J. Fluid Mech., 120:24-27) and evaluated numerically. The outcome is a surface-averaged value of membrane viscosity which is representative of a finite interval of membrane shear rate. The numerical values computed show a clear shear-thinning characteristic as well as a significant augmentation of viscosity with cell age and tend toward agreement with those determined for the rapid phase of shape recovery in micropipettes (Chien, S., K.-L. P. Sung, R. Skalak, S. Usami, and A. Tozeren, 1978, Biophys. J., 24:463-487). The computations also indicate that the rate of energy dissipation in the membrane is always substantially greater than that in the cytoplasm.

Viscosity of passive human neutrophils undergoing small deformations.

At issue is the type of constitutive equation that can be used to describe all possible types of deformation of the neutrophil. Here a neutrophil undergoing small deformations is studied by aspirating it into a glass pipet with a diameter that is only slightly smaller than the diameter of the spherically shaped cell. After being held in the pipet for at least seven seconds, the cell is rapidly expelled and allowed to recover its undeformed, spherical shape. The recovery takes approximately 15 s. An analysis of the recovery process that treats the cell as a simple Newtonian liquid drop with a constant cortical (surface) tension gives a value of 3.3 x 10(-5) cm/s for the ratio of the cortical tension to cytoplasmic viscosity. This value is about twice as large as a previously published value obtained with the same model from studies of large deformations of neutrophils. This discrepancy indicates that the cytoplasmic viscosity decreases as the amount of deformation decreases. An extrapolated value for the cytoplasmic viscosity at zero deformation is approximately 600 poise when a value for the cortical tension of 0.024 dyn/cm is assumed. Clearly the neutrophil does not behave like a simple Newtonian liquid drop in that small deformations are inherently different from large deformations. More complex models consisting either of two or more fluids or multiple shells must be developed. The complex structure inside the neutrophil is shown in scanning electron micrographs of osmotically burst cells and cells whose membrane has been dissolved away.

Hydrodynamics of a compound drop with application to leukocyte modeling

We study the dynamics of a compound liquid drop which is comprised of an outer membrane surface, a shell layer, and a core. The deformation due to an imposed extensional flow and the subsequent recovery are investigated computationally employing a combined Eulerian–Lagrangian technique. The numerical method allows for large viscosity and capillarity differences between layers. The present study reports several findings which provide direct insight into developing a dynamic model for leukocytes. A compound drop behaves like a homogeneous, simple liquid drop if the core is sufficiently deformed and the time scale of the core, related to the combination of its viscosity and capillarity, is comparable to that of the shell layer. Disparate time scales between the core and shell layer result in a rapid initial recoil of the drop during which the shell fluid is the primary participant in the hydrodynamics, followed by a slower relaxation period during which the core and shell layer interact with each other. Cons...

Effects of Nucleus on Leukocyte Recovery

AbstractThe rheological properties of a leukocyte significantly affect its biological and mechanical characteristics. To date, existing physical models of leukocyte are not capable of quantitatively explaining the wide range of deformation and recovery behaviors observed in experiment. However, a compound drop model has gained some success. In the present work, we investigate the effect of nucleus size and position, and the relative rheological properties of cytoplasm and nucleus, on cell recovery dynamics. Two nucleus sizes corresponding to that of neutrophil and lymphocyte are considered. Direct comparison between numerical simulations and experimental observation is made. Results indicate that the time scale ratio between the nucleus and cytoplasm plays an important role in cell recovery characteristics. Comparable time scales between the two cell components yield favorable agreement in recovery rates between numerical and experimental observations; disparate time scales, on the other hand, result in recovery behavior and cell shapes inconsistent with experiments. Furthermore, it is found that the nucleus eccentricity exhibits minimum influence on all major aspects of the cell recovery characteristics. The present work offers additional evidence in support of the compound cell model for predicting the rheological behavior of leukocytes.

Expression of GRP and Its Receptor in Well-differentiated Colon Cancer Cells Correlates with the Presence of Focal Adhesion Kinase Phosphorylated at Tyrosines 397 and 407

Gastrin-releasing peptide (GRP) and its receptor (GRP-R) are not normally expressed by epithelial cells lining the colon but are aberrantly expressed in cancer, where they act as morphogens and regulate tumor cell differentiation. Studies of colon cancer formation in mice genetically incapable of synthesizing GRP-R suggested that this receptors morphogenic properties were mediated via focal adhesion kinase (FAK). We therefore set out to determine the presence of both total and phosphorylated forms of FAK in human colon cancer specimens as a function of tumor cell differentiation and GRP/GRP-R co-expression. Ten colon cancers containing 25 regions of distinct differentiation were randomly selected from our GI Cancer Tumor Bank. All specimens were immunohistochemically probed using antibodies recognizing GRP, GRP-R, total FAK, and FAK specifically phos-phorylated at tyrosine (Y) 397, 407, 576, 577, 861, and 925. Antibody-specific chromogen was determined by quantitative immunohistochemistry (IHC) for each region of defined differentiation. Here we confirm that GRP/GRP-R co-expression is a function of differentiation, with highest levels observed in well-differentiated tumor cells. We also show that the amount of total FAK and of FAK phosphorylated at Y397 and Y407 tightly correlates with differentiation and with the amount of GRP/GRP-R co-expression. These findings are consistent with GRP/GRP-R acting as a morphogen by activating FAK, and suggest that this occurs via phosphorylation of this enzyme at two specific tyrosine residues.

A pneumatic vibrotactile stimulation device for fMRI

Mapping the functional response of the somatosensory cortex is useful both for characterizing normal brain activity and for determining the functional integrity of damaged cortex compromised by stroke or other neurological insults. A variety of stimulators have been used to produce somatosensory cortex activation in functional brain imaging, including brushes and swabs operated manually, pneumatically and mechanically powered mechanical vibrators, air puffs, and vibrating ceramic piezoelectric wafers and benders. A closed‐system, pneumatically driven rubber diaphragm is reported that overcomes many of the limitations of existing vibrotactile devices and produces robust sensory cortex activation in an fMRI experiment. Magn Reson Med 51:640–643, 2004.

Rheological modelling of leukocytes

A three-layer Newtonian model is investigated using a combined Eulerian-Lagrangian computational method to describe the dynamic behaviour of leukocytes. The model, composed of a cell membrane (outer layer), cytoplasm (middle layer) and nucleus (inner layer), can better describe the recovery characteristics because large viscosity and capillarity differences between layers are considered, and both Newtonian and seemingly non-Newtonian behaviours reported in the literature can be reproduced. It is found that, to describe adequately the various rheological characteristics of leukocytes, the presence of the highly viscous nucleus and its deformation/recovery, as well as the surface energy stored in the fluid interfaces, are critical. Photographs from pipette experiments using a fluorescent technique confirm the theoretical finding of the important role played by the nucleus in cell deformation.

Membrane stress and internal pressure in a red blood cell freely suspended in a shear flow

Presented is an algorithm for the approximate calculation of the membrane stress distribution and the internal pressure of a steadily tank-treading red cell. The algorithm is based on an idealized ellipsoidal model of the tank-treading cell (Keller, S.R., and R. Skalak, 1982, J. Fluid Mech., 120:27-47) joined with experimental observations of projected length, width, and tank-treading frequency. The results are inexact because the membrane shape and velocity are assumed a priori, rather than being determined via appropriate material constitutive relations for the membrane; these results are, nevertheless, believed to be approximately correct, and show that internal pressure builds up slowly as cell elongation increases, rising more rapidly as the deformed cell approaches the limiting geometry of a prolate ellipsoid. The maximum shear stress resultant in the membrane was found to be below but approaching the yield point range at the highest shear rate applied.

Phosphorylation of focal adhesion kinase tyrosine 397 critically mediates gastrin-releasing peptide's morphogenic properties.

We have proposed that gastrin‐releasing peptide (GRP) and its receptor (GRP‐R) are morphogens that when aberrantly re‐expressed in colon cancer promote tumor cell differentiation and retard metastasis. Because circumstantial evidence suggested that these properties were mediated via focal adhesion kinase (FAK), the purpose of this study was to elucidate the role of GRP‐induced activation of this enzyme on properties fundamental to metastasis including cell attachment, motility, and deformability. To do this, we studied 293 cells, a non‐malignant epithelial cell line that we show expresses GRP and GRPR. To dissect out the role of FAK, 293 cells were modified to inducibly express the dominant negative enzyme FAK‐related non‐kinase (FRNK) under control of a Tet‐On (i.e., doxycycline‐sensitive) promoter. Under serum‐free conditions, GRP acting in an autocrine manner caused FAK to be phosphorylated at Y397; and this could be completely inhibited either by incubating with the specific GRP‐R antagonist D‐Phe6(bombesin) methyl ester, or by upregulating FRNK using doxycycline. To measure cell attachment, we designed a cone‐plate viscometer that recorded the shear stress required to detach cells from their underlying matrix. To assess motility, confluent cells were wounded and behavior assessed by time‐lapse photography. To measure deformability, we recorded the ability of cells to be completely drawn into a micropipette <50% the size of the non‐deformed cell. Control 293 cells adhered more avidly to their underlying matrix, rapidly remodeled wounded tissues without any increase in overall proliferation, and were less distensible than cells treated with antagonist or doxycycline. Thus, these findings suggest that expression of GRP/GRPR in cancer inhibits metastasis by enhancing cell attachment to the matrix, regulating motility in the context of remodeling, and decreasing deformability. J. Cell. Physiol. 199: 77–88, 2004© 2003 Wiley‐Liss, Inc.