Quantifying rolling adhesion with a cell-free assay: E-selectin and its carbohydrate ligands.
Rolling of neutrophils over stimulated endothelial cells is a prerequisite to firm attachment and subsequent transendothelial migration during the inflammatory response. The selectin family of adhesion molecules are thought to mediate rolling by binding counter-receptors that present carbohydrates, such as sialyl Lewis(x) (sLe[x]). Recently we described a cell-free system for rolling using sLe(x)-coated microspheres and E-selectin molecules on inert substrates. We showed that sLe(x)-coated microspheres rolled over E-selectin-IgG chimera substrates with dynamics that are similar to those of leukocytes rolling over stimulated endothelium. In this paper we provide a thorough quantitative description of the dynamics of adhesion for this system. We find that particle rolling velocity increases with increasing wall shear stress and decreases with increasing E-selectin or sLe(x) surface densities. Large changes in the average rolling velocity can occur with small changes in sLe(x) or E-selectin density; however, rolling velocity is more sensitive to E-selectin surface coverage than to the number of sLe(x) molecules on the microspheres. Aided by dimensional analysis, we show that decreasing the wall shear stress or increasing either receptor (E-selectin) or ligand (sLe[x]) surface coverage results in an equivalent decrease in particle rolling velocity. In addition, we find that different Lewis carbohydrates are more effective in mediating rolling on E-selectin, with effectiveness following the trend sialyl Lewis(a) > sialyl Lewis(x) >> sulfated Lewis(x) >> Lewis(x). Rolling velocity fluctuated with time for all carbohydrate-selectin pairs tested, and the magnitude of the velocity fluctuations was linearly proportional to the mean rolling velocity for all combinations of E-selectin site density, sLe(x) site density, wall shear stress, and carbohydrate chemistry tested.
Brunk
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DK
School of Chemical Engineering, Cornell University, Ithaca, New York 14853, USA.
Hammer
D A
DA
D055592:Biophysical Phenomena; D001703:Biophysics; D050260:Carbohydrate Metabolism; D002448:Cell Adhesion / Q000502:physiology*; D002465:Cell Movement; D002474:Cell-Free System; D019040:E-Selectin / Q000502:physiology*; D004730:Endothelium, Vascular / Q000166:cytology; D005898:Glass; D006801:Humans; D007074:Immunoglobulin G; D066298:In Vitro Techniques; D008024:Ligands; D008863:Microspheres; D008954:Models, Biological; D009504:Neutrophils / Q000502:physiology; D009844:Oligosaccharides / Q000378:metabolism; D000080506:Sialyl Lewis X Antigen; D013499:Surface Properties
D019040:E-Selectin; D007074:Immunoglobulin G; D008024:Ligands; D009844:Oligosaccharides; D000080506:Sialyl Lewis X Antigen