Biomedical subjects

D A Hammer

Publications and source records attributed to D A Hammer.

50 records · Page 3Linked to original sources

PubMed1992

Simulation of cell rolling and adhesion on surfaces in shear flow: general results and analysis of selectin-mediated neutrophil adhesion.

The receptor-mediated adhesion of cells to ligand-coated surfaces in viscous shear flow is an important step in many physiological processes, such as the neutrophil-mediated inflammatory response, lymphocyte homing, and tumor cell metastasis. This paper describes a calculational method which simulates the interaction of a single cell with a ligand-coated surface under flow. The cell is idealized as a microvilli-coated hard sphere covered with adhesive springs. The distribution of microvilli on the cell surface, the distribution of receptors on microvilli tips, and the forward and reverse reaction between receptor and ligand are all simulated using random number sampling of appropriate probability functions. The velocity of the cell at each time step in the simulation results from a balance of hydrodynamic, colloidal and bonding forces; the bonding force is derived by summing the individual contributions of each receptor-ligand tether. The model can simulate the effect of many parameters on adhesion, such as the number of receptors on microvilli tips, the density of ligand, the rates of reaction between receptor and ligand, the stiffness of the resulting receptor-ligand springs, the response of springs to strain, and the magnitude of the bulk hydrodynamic stresses. The model can successfully recreate the entire range of expected and observed adhesive phenomena, from completely unencumbered motion, to rolling, to transient attachment, to firm adhesion. Also, the method can generate meaningful statistical measures of adhesion, including the mean and variance in velocity, rate constants for cell attachment and detachment, and the frequency of adhesion. We find a critical modulating parameter of adhesion is the fractional spring slippage, which relates the strain of a bond to its rate of breakage; the higher the slippage, the faster the breakage for the same strain. Our analysis of neutrophil adhesive behavior on selectin-coated (CD62-coated) surfaces in viscous shear flow reported by Lawrence and Springer (Lawrence, M.B., and T.A. Springer 1991. Cell. 65:859-874) shows the fractional spring slippage of the CD62-LECAM-1 bond is likely below 0.01. We conclude the unique ability of this selectin bond to cause neutrophil rolling under flow is a result of its unique response to strain. Furthermore, our model can successfully recreate data on neutrophil rolling as function of CD62 surface density.

Animals↗

PubMed1992

Equilibrium and kinetic analysis of Autographa californica nuclear polyhedrosis virus attachment to different insect cell lines.

The kinetic and equilibrium attachment of Autographa californica nuclear polyhedrosis virus (AcMNPV) to seven insect cell lines was evaluated. Kinetic experiments revealed differences of up to 10-fold in the infection rates among cell lines. Equilibrium binding also varied between cell lines and was saturable. The Tn 5B1-4 and Tn F cell lines had the highest virus binding affinities and infection rates and exhibited diffusion-limited attachment. The rate of infection appears to be limited by the rate of attachment. For the Tn 5B1-4 cells the physical to infective particle ratio for AcMNPV was 5.3. From the Scatchard analyses, the cell lines Tn 5B1-4 and Tn F displayed affinities of 2.35 x 10(10) M-1 and 1.60 x 10(10) M-1, respectively, with 6000 and 13,700 binding sites per cell. The insect cell line Hz 1075, which is not susceptible to AcMNPV infection, displayed a much lower, but saturable, binding of AcMNPV with 900 sites/cell and an affinity of 1.1 x 10(10) M-1. Unlabelled AcMNPV, but not Lymantria dispar MNPV could compete with labelled AcMNPV for binding sites. There were 93 to 96% reductions in virus cell binding following pretreatments of cells with three proteases, suggesting the involvement of a cellular protein component in virus binding. Tunicamycin, an inhibitor of N-linked glycosylation and expression of some membrane proteins on the cell surface, reduced virus binding in a dose-dependent manner suggesting a role for glycoprotein(s) in binding. However there was no evidence for the direct involvement of oligosaccharides in attachment. Metabolic inhibitors of oligosaccharide trimming and competition binding assays using simple sugars caused no measurable reductions in virus binding. These findings suggest that AcMNPV attachment to insect cells is receptor-mediated via a glycoprotein component(s); the direct involvement of oligosaccharide moieties in binding is unlikely.

Animals↗

PubMed1991

Simulation of cell rolling and adhesion on surfaces in shear flow. Microvilli-coated hard spheres with adhesive springs.

The adhesion of cells to ligand-coated surfaces in viscous shear flow is an important step in many physiological processes, such as the neutrophil-mediated inflammatory response, lymphocyte homing, and tumor cell metastasis. This article describes a calculational method that allows simulation of the interaction of a single cell with a ligand-coated surface. The cell is idealized as a microvilli-coated hard sphere covered with adhesive springs. The distribution of microvilli on the cell surface, the distribution of receptors on microvilli tips, and the forward and reverse reaction between receptor and ligand are all simulated using random number sampling of appropriate probability functions. The velocity of the cell at each time step in the simulation results from a balance of hydrodynamic, colloidal, and bonding forces; the bonding force is derived by summing the individual contributions of each receptor-ligand tether. The model can simulate the effect of many parameters on adhesion, such as the number of receptors on microvilli tips, the density of ligand, the rates of reaction between receptor and ligand, the stiffness of the springs, the response of springs to extension, and the magnitude of hydrodynamic stresses. By varying these parameters, the model can successfully recreate the entire range of expected and observed adhesive phenomena, from completely unencumbered motion, to rolling, to transient attachment, to firm adhesion. Also, the model can provide meaningful statistical measures of adhesion, including the mean and variance in velocity, rate constants for cell attachment and detachment, and the frequency of adhesion. We find a critical modulating parameter of adhesion is the fractional spring slippage, which relates the extension of a bond to its rate of breakage; the higher the slippage, the faster the breakage for the same extension. Changes in the fractional spring slippage can radically change the adhesive behavior of a cell. We show that stiffer springs will only serve to increase adhesion if the fractional slippage remains small. In addition, our simulations emphasize the importance of reaction rates between receptor and ligand, rather than affinity, as being the key determinant of adhesion under flow. These results suggest reaction rates and response to stress of adhesion molecules must be independently measured to understand how adhesion is controlled at the molecular level.

Animals↗

PubMed1990

Organ-preference of metastasis. The role of endothelial cell adhesion molecules.

The initial, site-specific colonization of secondary organs by blood-borne cancer cells appears to be mediated by endothelial cell adhesion molecules. These molecules are part of the organ-specific microvascular phenotype and are regulated through complex interactions of the endothelium with the extracellular matrix (e.g., distinct matrix macromolecules and growth factors). They are induced in vitro by growing 'unspecific' (large vessel) endothelial cells on extracts of organ-specific biomatrices. In many respects, these molecules are similar to the various classes of chemically different adhesion molecules that regulate lymphocyte traffic, but are believed to be distinct from the inducible adhesion molecules that govern leukocyte adhesion during acute episodes of inflammation. Biochemical and biophysical data indicate that preference of tumor cell adhesion to organ-specific microvascular endothelium may not require qualitative differences of such homing receptors between endothelia, but may be explained on the basis of quantitative receptor differences as well as differences of receptor avidity. Following adhesion, the metastatic cascade proceeds by the establishment of metabolic conduits between the endothelium and adherent tumor cells. This heterotypic coupling represents an early step in the extravasation of cancer cells from the microvasculature, initiating endothelial cell retraction from its basement membrane and recanalization around the arrested tumor cell. These events, together with local growth promoting effects exerted by the metastasized organ, are believed to provide the basis for Paget's 'seed and soil' hypothesis of metastasis.

Animals↗

PubMed1990

General analysis of receptor-mediated viral attachment to cell surfaces.

Viruses are multivalent particles that attach to cells through one or more bonds between viral attachment proteins (VAP) and specific cellular receptors. Three modes of virus binding are presented that can explain the diversity in binding data observed among viruses. They are based on multivalency of attachment and spatial versus receptor saturation effects which are easily distinguished based upon simple criteria. Mode 1 involves only monovalent virus/receptor binding. Modes 2 and 3 involve multivalent bonds between the virus and cell; however, in mode 3 space on the cell surface becomes saturated before receptors. A model is developed for viral attachment that accounts for nonspecific binding, receptor/virus interactions, and spatial saturation effects. The model can describe each mode in different limits and can be applied to virus binding data to extract key physical information such as receptor number and affinity. These values are used to postulate the type of VAP/receptor interaction involved and to predict binding at different parameter values. For the mode 2 binding of Adenovirus 2, the model predicts a receptor number of 4-15 x 10(3) on HeLa cells and an affinity of 2-6 x 10(7) M-1 which closely approximate experimental estimates. For the binding of three, broad-host-range, enveloped viruses, Semliki Forest virus, Vesicular Stomatitis virus, and the baculovirus, Autographa californica nuclear polyhedrosis virus, the model predicts receptor numbers of 10(5) or greater and affinities in the range of 10(4) to 10(5) M-1. These values are indicative of a VAP/oligosaccharide interaction which has been documented for a number of other viruses. Experimental evidence is presented that is the first to demonstrate that baculovirus binding is mediated by a cell surface receptor.

Animals↗

PubMed1990

Bioreactor development for production of viral pesticides or heterologous proteins in insect cell cultures.

The insect cell-baculovirus expression system has significant potential for producing proteins requiring some degree of posttranslational modification. T. ni cells appear to be as good a host as S. frugiperda cells for heterologous protein production as demonstrated by production of beta-galactosidase. Attachment-dependent cells of T. ni can be effectively cultured in a packed-bed reactor using glass beads. When cell in such a reactor were infected, they produced 35% of the total protein as beta-galactosidase. No cell detachment was observed even 70 h postinfection. A model of viral entry has been proposed and tested.

Animals↗

PubMed1989

A dynamical model for receptor-mediated cell adhesion to surfaces in viscous shear flow.

We present a dynamical model for receptor-mediated cell adhesion to surfaces in viscous shear flow when the surfaces are coated with ligand molecules complementary to receptors in the cell membrane. This model considers the contact area between the cell and the surface to be a small, homogeneous region that mediates the initial attachment of the cell to the surface. Using a phase plane analysis for a system of nonlinear ordinary differential equations that govern the changes in free receptor density and bond density within the contact area with time, we can predict the conditions for which adhesion between the cell and the surface will take place. Whether adhesion occurs depends on values of dimensionless quantities that characterize the interaction of the cell and its receptors with the surface and its ligand, such as the bond formation rate, the receptor-ligand affinity, the fluid mechanical force, the receptor mobility, and the contact area. A key result is that there are two regimes in which different chemical and physical forces dominate: a rate-controlled high affinity regime and an affinity-controlled low affinity regime. Many experimental observations, including the effects of temperature and receptor mobility on adhesiveness, can be explained by understanding which of these regimes is appropriate. We also provide simple approximate analytical solutions, relating adhesiveness to cell and surface properties as well as fluid forces, which allow convenient testing of model predictions by experiment.

Animals↗

PubMed1987

A dynamical model for receptor-mediated cell adhesion to surfaces.

We present a dynamical model for receptor-mediated adhesion of cells in a shear field of viscous fluid to surfaces coated with ligand molecules complementary to receptors in the cell membrane. We refer to this model as the "point attachment model" because it considers the contact area between the cell and the surface to be a small, homogeneous region that mediates the initial attachment of the cell to the surface. Using a phase plane analysis of a system of nonlinear ordinary differential equations which govern the changes in free receptor density and bond density within the contact area with time, we can predict the conditions for which adhesion between the cell and the surface will take place. Whether adhesion occurs depends on values of dimensionless quantities that characterize the interaction of the cell and its receptors with the surface and its ligand, such as the bond formation rate, the receptor-ligand affinity, the fluid mechanical force, the receptor mobility, and the contact area. A key result is that there are two regimes in which different chemical and physical forces dominate: a rate-controlled high affinity regime and an affinity-controlled low-affinity regime. Many experimental observations can be explained by understanding which of these regimes is appropriate. We also provide simple approximate analytical solutions, relating adhesiveness to cell and surface properties as well as fluid forces, which allow convenient testing of model predictions by experiment.

Animals↗

PubMedDate not supplied

Kinetics of cell detachment: effect of ligand density.

Cell adhesion to substratum is often mediated by binding between cell surface receptors and substrate ligands. Substrates can be derivatized with different types and densities of ligands, but how substrate chemistry determines cellular function, such as adhesion strength, has not been demonstrated quantitatively. We employ a numerical methodology developed by Dembo and colleagues (9), who investigated membrane peeling under conditions of excess ligand density, to investigate the kinetics and strength of cell peeling from ligand coated surfaces for arbitrary ligand density. We show there are two asymptotic limits to peeling strength, as quantified by the critical tension: a high ligand density limit, where the critical tension is independent of ligand density and depends logarithmically on the receptor density; and a low ligand density limit, in which the critical tension depends logarithmically on the ligand density but is independent of receptor density. In between these limits, we numerically determine the critical tension. The critical tension is always a weak function of the dissociation constant between ligand and receptor. Furthermore, we show how the rate of peeling, for tensions above the critical tension, depends on ligand density and the mechanical properties of the receptor-ligand bonds. Interestingly, we illustrate when small increases in ligand density should alter cellular behavior, inducing a change to spreading onto a substrate from peeling up from a substrate. In total the predictions of this paper provide criteria for the design of ligand-coated substrate that provide for the proper adhesion strength and dynamics of detachment of cells from surfaces.

Animals↗

PubMedDate not supplied

Rheology of rat basophilic leukemia cells.

Rat basophilic leukemia (RBL) cells, decorated with IgE, have been shown to bind irreversibly to antigen-coated substrates. In this paper we measured RBL cell deformability and demonstrated that this irreversible binding is not due to a compliant cellular rheology of these cells. The rheological properties of RBL cells were assessed with single-cell micropipette aspiration. Small-sized (G1/G0 phase) cells were found to be more deformable than medium-sized (S phase) cells. No changes in cellular rheology were observed after binding of anti-dinitrophenol IgE to Fce receptors. Furthermore, cytoplasmic viscosity mu showed power-law dependence on mean shear rate gama m: mu = mu c(gamma m/gamma c)-b, where mu c is a characteristic viscosity at characteristic shear rate gamma c, and b is a material coefficient. All the cells exhibited similar dependence on shear rate (b approximately 0.5). When gamma c was set to 1 s-1. mu c = 480 +/- 10,560 +/- 40 and 490 +/- 10 Pa.s for G1/G0, S cells, and G1/G0 cells treated with the antibody, respectively. In general. RBL cells were much more rigid than normal neutrophils (mu c = 130 +/- 20 Pa, s b = 0.5). Thus the biochemistry of the adhesion molecules, not the cellular deformability of the cell, is the cause of the irreversibility of RBL cell adhesion under flow.

2,4-Dinitrophenol↗

PubMedDate not supplied

An E-selectin-IgG chimera supports sialylated moiety dependent adhesion of colon carcinoma cells under fluid flow.

To further characterize the molecular mechanisms that govern carcinoma cell adhesion to stimulated endothelium, we studied the adhesion of a human colon carcinoma cell line, KM12-L4, to an E-selectin-IgG1 chimera and interleukin (IL)-1 beta-stimulated human umbilical vein endothelial cells (HUVEC) under in vitro fluid flow conditions. Between 0.6 and 1.8 dynes/cm2, KM12-L4 cells attach to and roll on IL-1 beta-stimulated HUVEC. The adhesion is E-selectin dependent and diminished upon pretreatment of the KM12-L4 cells with neuraminidase (neuraminidase sensitive). Between 0.7 and 1.8 dynes/cm2, surfaces coated with an E-selectin-IgG1 chimera support attachment and rolling of KM12-L4 cells. The adhesion to the E-selectin-IgG1 chimera is blocked by an antibody to the lectin domain of E-selectin and is neuraminidase sensitive. Rolling KM12-L4 cells exhibit variable velocity motion over both IL-1 beta-stimulated HUVEC and E-selectin-IgG1 chimera-coated surfaces. Our results provide the first direct evidence that sialylated moieties are involved in the adhesion of carcinoma cells to IL-1 beta-stimulated endothelium under flow conditions; E-selectin-IgG1 chimeras can support cell attachment and rolling under defined flow conditions; the topology of the endothelium is not the sole cause of variable velocity motion observed in cell rolling systems.

Cell Adhesion↗

PubMedDate not supplied

Morphology of cell-substratum adhesion. Influence of receptor heterogeneity and nonspecific forces.

Many cell types modulate growth, differentiation, and motility through changes in cell substrate adhesion, including regulation of focal contact formation. Clustering of cell surface adhesion receptors is an essential early step in the development of focal contacts, and thus may influence cell physiology. In this paper, we present a theoretical framework to examine how cell surface chemistry affects receptor clustering. Our one-dimensional tape-peeling model couples the equations of mechanical equilibrium for a cell membrane with kinetic receptor-ligand binding relations. We considered two distinct model scenarios: Adhesion mediated by multiple receptor-ligand interactions of different length and specific binding of a single receptor type occurs in the presence of van der Waals attraction and nonspecific repulsion. In each case, nonuniform (wave-like) membrane morphologies are observed in certain parameter ranges that support the clustering of adhesion receptors. The formation of these morphologies is described in terms of a balance of membrane stresses; when cell-surface potential as a function of separation distance is symmetric between two potential energy minima, nonuniform morphologies are obtained. Increases in the chemical binding energy between receptor and ligand (e.g., increases in ligand density) or decreases in the membrane rigidity result in smaller wavelengths for nonuniform interfaces. Additionally, we show wave-like geometries appear only when the mechanical compliance of receptor-ligand bonds is within an intermediate range, and examine how the mobility of "repellers"--glycocalyx molecules that exert a nonspecific repulsive force--influences membrane morphology. We find fully mobile repellers always redistribute to prevent nonuniform morphologies.

Animals↗

PubMedDate not supplied

Motion of model leukocytes near a wall in simple shear flow.

Receptor-mediated cell adhesion to surfaces depends on the motion of the cell prior to adhesion. We studied the hydrodynamic behavior of cells near a wall in simple shear flow using a model leukocyte system that we have also used extensively in cell-substrate adhesion studies. Specifically, we measured the velocity of rat basophilic leukemia (RBL) cells near a surface in a parallel-plate flow chamber and compared it to the motion of polystyrene beads and glutaraldehyde-fixed RBL cells. We found that RBL cells (13 microns diameter) travel 25% faster than polystyrene beads of 14.5 microns diameter for a wide range of shear rates (20-180 s-1); this suggests that RBL cells would travel 39% faster than polystyrene beads of equivalent size. Glutaraldehyde-fixed RBL cells travel at a velocity between those of live cells and 14.5-microns beads. These differences in velocities have been observed over both polyacrylamide gel and glass substrates. Application of a theory for hard sphere motion near a wall in simple shear flow at low Reynolds number [Goldman, A.J.; Cox, R.G.; Brenner, H. Chem. Eng. Sci. 1967b, 22, 653-660] to our measured cell velocities suggests that cells are separated from the wall by > or = 550 nm. Such large separation distances have also been predicted by others who have used hard sphere theory to analyze the effect of shear flow on cell motion near walls. However, the extensive receptor-mediated cell adhesion seen in these systems is inconsistent with these separation distances, which are approximately 30 times greater than the distance required for receptor-ligand contact. Instead, we propose that, because of cell deformability and cell surface roughness, cells remain within a separation distance that allows for molecular contact while they travel faster than the hard sphere theory predicts. Therefore, the theory of Goldman and co-workers, while adequate for hard sphere motion, is likely not accurate for cellular motion.

Animals↗

PubMedDate not supplied

A simple model to predict the effectiveness of molecules that block attachment of human rhinoviruses and other viruses.

The binding of viruses to cell surfaces is often mediated by cell surface receptors. The use of soluble receptors, such as intracellular adhesion molecule-1 (ICAM-1) for human rhinovirus (HRV), CD4 for human immunodeficiency virus (HIV), and CR2 for Epstein-Barr virus, for in vivo antiviral therapy is under serious investigation. A number of synthetic compounds that affect HRV attachment and uncoating (termed WIN compounds) are also being studied. However, the mechanism behind the dose-response effect of these agents in preventing infection has not been clearly demonstrated. In addition to simple blocking (by receptors) or inactivation of binding sites (by WIN compounds) on the virus surface, other mechanisms of inhibition have been proposed and demonstrated, including cooperative inactivation of neighboring sites, receptor-induced viral attachment protein (VAP) shedding, virus particle inactivation, and inhibition of multivalent virus binding. We present a simple mathematical model to predict the effect of these molecules on virus infection by incorporating only the blocking or site inactivation step of the blocking molecule and its resulting effect on attachment. The ability of the model to reproduce the response of a virus to a dose of blocking molecules is used to distinguish the role of blockers in inhibiting attachment from the other mechanisms of viral inactivation that have been proposed. The model includes both the reversible attachment of the virus to its cellular receptor and to soluble receptors or synthetic molecules (blockers).(ABSTRACT TRUNCATED AT 250 WORDS)

Antiviral Agents↗