Immobilized IL-8 triggers progressive activation of neutrophils rolling invitro on P-selectin and intercellular adhesion molecule-1

The chemokine IL-8 is found on the luminal side of vascular endothelial cel ls, where it is postulated to be immobilized during inflammation. In this s tudy, we observed that immobilized IL-8 can stimulate neutrophils to firmly adhere to a substrate containing ICAM-1 in a static adhesion assay. Solubl e IL-8 was then perfused over neutrophils rolling on P-selectin (P-sel) and ICAM-1, confirming that IL-8 in solution can quickly cause rolling neutrop hils to arrest. To mimic a blood vessel wall with IL-8 expressed on the lum inal surface of endothelial cells, IL-8 was immobilized along with P-sel an d ICAM-1 at defined site densities to a surface. Neutrophils rolled an aver age of 200 mum on surfaces of P-sel, ICAM-1, and IL-8 before firmly adherin g through ICAM-1-beta (2) integrin interactions at 2 dynes/cm(2) wall shear stress. Increasing the density of IL-8 from 60 to 350 sites/mum(2) on the surface decreased by 50% the average distance and time the neutrophils roll ed before becoming firmly adherent. Temporal dynamics of ICAM-1-beta (2) in tegrin interactions of rolling neutrophils following IL-8 exposure suggest the existence of two classes of beta (2) integrin-ICAM-1 interactions, a lo w avidity interaction with a 65% increase in pause times as compared with P -sel-P-sel glycoprotein ligand-1 interactions, and a high avidity interacti on with pause times 400% greater than the selectin interactions. Based on t he proportionality between IL-8 site density and time to arrest, it appears that neutrophils may need to sample a critical number of IL-8 molecules pr esented by the vessel wall before forming a sufficient number of high avidi ty beta (2) integrin bonds for firm adhesion.