Chemokines are widely hypothesized to stimulate firm adhesion of leukocytes on endothelium in shear flow. Thus far, this has been demonstrated experimentally for exogenously added chemoattractants, but not for those released by endothelium. We found that human umbilical cord endothelial cells (HUVEC) stimulated with TNF-α and IFN-γ secreted eosinophil chemoattractants into the culture supernatant. This material induced transendothelial chemotaxis, stimulated eosinophil binding to purified intercellular adhesion molecule 1, and augmented binding to purified vascular cell adhesion molecule 1 in a 3-min static assay. Chemotaxis and stimulation of adhesion were abrogated completely by the pretreatment of eosinophils with an mAb to the C-C chemokine receptor 3 (CCR3). Eosinophils accumulated efficiently on HUVEC stimulated with TNF-α and IFN-γ in shear flow at 1.5 dyn/cm2. CCR3 mAb slightly but significantly reduced eosinophil arrest and accumulation, by preventing development of firm adhesion by some of the tethered eosinophils, so that they detached within 30 s after the initial tethering. In the presence of mAb to the α4 integrin subunit, the effect of CCR3 mAb was more prominent, and approximately half of eosinophil arrest and accumulation was abolished. Inhibition by CCR3 mAb in the presence of β2 integrin mAb was similar to that in control eosinophils. This is the first evidence that endothelial cell-derived chemokines can activate firm adhesion through α4 and β2 integrins even in the presence of shear flow.
- Chemokine receptor
- Very late antigen 4