Adhesive dynamics simulation of neutrophil arrest with stochastic activation.

The transition from rolling to firm adhesion is a key step in the adhesion cascade that permits a neutrophil to exit the bloodstream and make its way to a site of inflammation. In this work, we construct an integrated model of neutrophil activation and arrest that combines a biomechanical model of neutrophil adhesion and adhesive dynamics, with fully stochastic signal transduction modeling, in the form of kinetic Monte Carlo simulation within the microvilli. We employ molecular binding parameters gleaned from the literature and from simulation of cell-free rolling mediated by selectin molecules. We create a simplified model of lymphocyte function-associated antigen-1 activation that links P-selectin glycoprotein ligand-1 ligation to integrin activation. The model utilizes an energy profile of various integrin activation states drawn from literature data and permits manipulation of signal diffusivity within the microvillus. Our integrated model recreates neutrophil arrest within physiological timescales, and we demonstrate that increasing signal diffusivity within a microvillus accelerates arrest. If the energy barrier between free unactivated and free activated lymphocyte function-associated antigen-1 increases, the period of rolling before arrest increases. We further demonstrate that, within our model, modification of endothelial ligand surface densities can control arrest. In addition, the relative concentrations of signaling molecules control the fractional activation of the overall signaling pathway and the rolling time to arrest. This work presents the first, to our knowledge, fully stochastic model of neutrophil activation, which, though simplified, can recapitulate significant physiological details of neutrophil arrest yet retains the capacity to incorporate additional information regarding mechanisms of neutrophil signal transduction as they are elucidated.

Adhesive dynamics simulation of neutrophil arrest with deterministic activation.

The transition from rolling to firm adhesion is a key element of neutrophil activation and essential to the inflammatory response. Although the molecular mediators of rolling and firm adhesion are known to be selectins and beta2 -integrins, respectively, the precise dynamic mechanism by which these ligands facilitate neutrophil arrest remains unknown. Recently, it has been shown that ligation of E-selectin can stimulate the firm adhesion of neutrophils via a MAP-kinase cascade. To study the possible mechanism by which neutrophil arrest could occur, we created an integrated model by combining two methodologies from computational biology: a mechanics-based modeling of leukocyte adhesion (adhesive dynamics) and signal transduction pathway modeling. Within adhesive dynamics, a computational method our group has shown to accurately recreate rolling dynamics, we include a generic, tunable integrin activation module that links selectin engagement to integrin and activity. This model allows us to relate properties of the activation function to the dynamics of rolling and the time and distance rolled before arrest. This integrated model allows us to understand how intracellular signaling activity can set the timescale of neutrophil activation, adhesion, and diapedesis.

I-domain of lymphocyte function-associated antigen-1 mediates rolling of polystyrene particles on ICAM-1 under flow.

In their active state, beta(2)-integrins, such as LFA-1, mediate the firm arrest of leukocytes by binding intercellular adhesion molecules (ICAMs) expressed on endothelium. Although the primary function of LFA-1 is assumed to be the ability to mediate firm adhesion, recent work has shown that LFA-1 can contribute to cell tethering and rolling under hydrodynamic flow, a role previously largely attributed to the selectins. The inserted (I) domain of LFA-1 has recently been crystallized in the wild-type (wt) and locked-open conformations and has been shown to, respectively, support rolling and firm adhesion under flow when expressed in alpha(L)beta(2) heterodimers or as isolated domains on cells. Here, we report results from cell-free adhesion assays where wt I-domain-coated polystyrene particles were allowed to interact with ICAM-1-coated surfaces in shear flow. We show that wt I-domain can independently mediate the capture of particles from flow and support their rolling on ICAM-1 surfaces in a manner similar to how carbohydrate-selectin interactions mediate rolling. Adhesion is specific and blocked by appropriate antibodies. We also show that the rolling velocity of I-domain-coated particles depends on the wall shear stress in flow chamber, I-domain site density on microsphere surfaces, and ICAM-1 site density on substrate surfaces. Furthermore, we show that rolling is less sensitive to wall shear stress and ICAM-1 substrate density at high density of I-domain on the microsphere surface. Computer simulations using adhesive dynamics can recreate bead rolling dynamics and show that the mechanochemical properties of ICAM-1-I-domain interactions are similar to those of carbohydrate-selectin interactions. Understanding the biophysics of adhesion mediated by the I-domain of LFA-1 can elucidate the complex roles this integrin plays in leukocyte adhesion in inflammation.

A semianalytic model of leukocyte rolling.

Rolling allows leukocytes to maintain adhesion to vascular endothelium and to molecularly coated surfaces in flow chambers. Using insights from adhesive dynamics, a computational method for simulating leukocyte rolling and firm adhesion, we have developed a semianalytic model for the steady-state rolling of a leukocyte. After formation in a force-free region of the contact zone, receptor-ligand bonds are transported into the trailing edge of the contact zone. Rolling velocity results from a balance of the convective flux of bonds and the rate of dissociation at the back edge of the contact zone. We compare the model's results to that of adhesive dynamics and to experimental data on the rolling of leukocytes, with good agreement. We calculate the dependence of rolling velocity on shear rate, intrinsic forward and reverse reaction rates, bond stiffness, and reactive compliance, and use the model to calculate a state diagram relating molecular parameters and the dynamic state of adhesion. A dimensionless form of the analytic model permits exploration of the parameters that control rolling. The chemical affinity of a receptor-ligand pair does not uniquely determine rolling velocity. We elucidate a fundamental relationship between off-rate, ligand density, and reactive compliance at the transition between firm and rolling adhesion. The model provides a rapid method for screening system parameters for the potential to mediate rolling.

Polyethylene glycol additives reduce hemolysis in red blood cell suspensions exposed to mechanical stress.

Mechanical damage to blood cells is of considerable concern in the development and use of circulatory assist devices and other blood contacting systems. Furthermore, hemodilution with saline, dextran, and other plasma expanders applied during extracorporeal circulation and dialysis increases red blood cell (RBC) susceptibility to the high shear stresses associated with these procedures. In this paper, we present polyethylene glycol (PEG) as a potential erythrocyte protective agent against mechanically induced cellular trauma. Bovine RBCs were subjected to mechanical stress induced by rolling stainless steel shots through RBC suspensions for a constant exposure time. The suspensions were prepared at a hematocrit of 30% in various media: PEG (20,000 molecular weight), autologous bovine plasma, Dextran 40 solution, and phosphate buffered saline (PBS). RBC suspensions in Dextran 40 were prepared at a viscosity similar to the PEG suspensions. We found the hemolysis level of RBCs suspended in plasma and in PEG solutions to be several times lower (p < 0.001) than in the Dextran and PBS solutions. No statistically significant difference was found between the hemolysis that occurred in suspensions of RBCs in autologous plasma and in 2.0% PEG solutions. Even PEG concentration as low as 0.1% reduced hemolysis by more than 40% compared with PBS or the same concentration of Dextran in suspension medium. Our data demonstrate the efficacy of PEG molecules in reducing mechanical trauma to erythrocytes and suggest the potential for using PEG in assisted circulation, dialysis, and other procedures where RBCs are subjected to extensive mechanical stress.