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Biophys J, November 2000, p. 2391-2402, Vol. 79, No. 5

Cell-Free Rolling Mediated by L-Selectin and Sialyl Lewisx Reveals the Shear Threshold Effect

Adam W. Greenberg, Debra K. Brunk, and Daniel A. Hammer

Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104 USA

The selectin family of adhesion molecules mediates attachment and rolling of neutrophils to stimulated endothelial cells. This step of the inflammatory response is a prerequisite to firm attachment and extravasation. We have reported that microspheres coated with sialyl Lewisx (sLex) interact specifically and roll over E-selectin and P-selectin substrates (Brunk et al., 1996; Rodgers et al., 2000). This paper extends the use of the cell-free system to the study of the interactions between L-selectin and sLex under flow. We find that sLex microspheres specifically interact with and roll on L-selectin substrates. Rolling velocity increases with wall shear stress and decreases with increasing L-selectin density. Rolling velocities are fast, between 25 and 225 µm/s, typical of L-selectin interactions. The variability of rolling velocity, quantified by the variance in rolling velocity, scales linearly with rolling velocity. Rolling flux varies with both wall shear stress and L-selectin site density. At a density of L-selectin of 800 sites/µm2, the rolling flux of sLex coated microspheres goes through a clear maximum with respect to shear stress at 0.7 dyne/cm2. This behavior, in which the maintenance and promotion of rolling interactions on selectins requires shear stress above a threshold value, is known as the shear threshold effect. We found that the magnitude of the effect is greatest at an L-selectin density of 800 sites/µm2 and gradually diminishes with increasing L-selectin site density. Our study is the first to reveal the shear threshold effect with a cell free system and the first to show the dependence of the shear threshold effect on L-selectin site density in a reconstituted system. Our ability to recreate the shear threshold effect in a cell-free system strongly suggests the origin of the effect is in the physical chemistry of L-selectin interaction with its ligand, and largely eliminates cellular features such as deformability or topography as its cause.

Biophys J, November 2000, p. 2391-2402, Vol. 79, No. 5
© 2000 by the Biophysical Society   0006-3495/00/11/2391/12  $2.00


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