Biomechanics of P-selectin PSGL-1 bonds: Shear threshold and integrin-independent cell adhesion

¹ State University of New York at Buffalo
² McGill University
³ Pacific Northwest National Laboratory

^* To whom correspondence should be addressed. E-mail: neel{at}eng.buffalo.edu.

Submitted on May 4, 2005
Revised on June 24, 2005
Accepted on 5 December 2005

	Abstract

Platelet-leukocyte adhesion may contribute to thrombosis and inflammation. We examined the heterotypic interaction between unactivated neutrophils and either thrombin receptor activating peptide (TRAP) stimulated platelets or P-selectin bearing beads (Ps-beads) in suspension. Cone-plate viscometers were used to apply controlled shear rates from 14-3000/s. Platelet-neutrophil and bead-neutrophil adhesion analysis was performed using both flow cytometry and high-speed videomicroscopy. We observed that while blocking antibodies against either P-selectin or P-selectin glycoprotein ligand-1 (PSGL-1) alone inhibited platelet-neutrophil adhesion by ~60% at 140/s, these reagents completely blocked adhesion at 3000/s. Anti-Mac-1 alone did not alter platelet-neutrophil adhesion rates at any shear rate, though in synergy with selectin antagonists it abrogated cell binding. Unstimulated neutrophils avidly bound Ps-beads and activated platelets in an integrin-independent manner, suggesting that purely selectin-dependent cell adhesion is possible. In support of this, antagonists against P-selectin or PSGL-1 dissociated previously formed platelet-neutrophil and Ps-bead neutrophil aggregates under shear in a variety of experimental systems, including in assays performed with whole blood. In studies where medium viscosity and shear rate were varied, a shear threshold for P-selectin PSGL-1 binding was also noted at shear rates<100/s when Ps-beads collided with isolated neutrophils. Results are discussed in light of biophysical computations that characterize the collision between unequal size particles in linear shear flow. Overall, our studies reveal an integrin-independent regime for cell adhesion and weak shear-threshold for P-selectin PSGL-1 interactions that may be physiologically relevant.

Key Words: bond lifetime, flow cytometry, high-speed videomicroscopy, leukocyte, neutrophil, platelet

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