Leukocyte adhesion kinetics: A sticky business

Leukocyte rolling during inflammation is mediated primarily by the selectin family of adhesion molecules. P-selectin glycoprotein ligand-1 (PSGL-1) is a highly characterized molecule and is the primary ligand for P-selectin. PSGL-1 is a dimeric molecule with the two identical chains linked by a disulfide bond. I have measured the contributions of dimerization and glycosylation, in the binding pocket of PSGL-1, to bonds formed during rolling and tethering. I have also used a Monte Carlo simulation to model the idealized version of leukocyte rolling, which is the rolling of microbeads with adsorbed PSGL-1 on an adsorbed P-selectin surface.; The PSGL-1 isolated from transfected cell lines had different molecular weights depending on the activity of the core-2 glucosaminyltransferase. The high molecular weight species of PSGL-1 migrated a distance corresponding to ∼140 kD, consistent with PSGL-1 from a human leukemia cell line HL60, and the low molecular weight species of PSGL-1 migrated a distance corresponding to ∼120 kD under reducing conditions. The lack of core-2 activity in specific cell lines apparently led to a deficiency in PSGL-1---P-selectin bond strength. Cells with the low molecular weight species of PSGL-1 also formed tethers with a P-selectin substrate less frequently and rolled an order of magnitude faster than cells with the high molecular weight species. Cellular tethers and bonds formed with the low molecular weight species of PSGL-1 were more compliant as the dissociation rate constant increased rapidly with increased force. The high molecular weight species of PSGL-1 conferred a strengthening of bonds as the dissociation rate constants increased less with force, similar to that seen with PSGL-1 tethers from neutrophils.; Dimerization of PSGL-1 did not affect rolling velocity as cells expressing both monomeric and dimeric PSGL-1 rolled 10 mum/s or less, which is similar to neutrophil rolling velocities at the same conditions. The distance between tethering events was also unaffected by dimerization. The initial tethering from flow or capture, when the cell first initiates contact with the P-selectin substrate, did increase with dimerization over the accumulation of tethering cells expressing the monomeric PSGL-1. The bond strength or variation of dissociation rate with force did not depend on PSGL-1 dimerization, implying that a tether mediated by dimeric PSGL-1 is the same as a tether mediated by monomeric PSGL-1 and they consist of similar numbers of bonds.; The Monte Carlo simulation of microbead rolling matched experimental values of microbead rolling velocity at several different wall shear stresses. The case of the model implementing multiple tethers or bonds was able to match the experimental values for all wall shear stresses, but varied in separation distance between the bead surface and the substrate. Up to two tethers could form, in the case of multiple tethers. Re-bonding, once one tether broke and the other remained, was also possible. This model matched experimentally measured bead rolling velocities at higher separation distances for higher wall shear stresses. The results from the simulation imply that multiple tethers are required for bead rolling for the conditions modeled and a hydrodynamic lift force may be affecting bead separation distance even during rolling.