Neutrophil adherence to vascular endothelial cells is the initial event in the emigration of neutrophils through blood vessel walls to tissue sites of inflammation; this process is attributed to the generation of extravascular chemotactic factors. To investigate the effect of chemotactic factors on neutrophil adherence to endothelium, we developed a sensitive, reproducible in vitro microtiter adherence assay. Base-line nonstimulated adhesion of human neutrophils to cultured human umbilical vein endothelial cell monolayers was 35.2 +/- 0.9%, which is equivalent to three to four neutrophils per endothelial cell. Addition of either purified complement fragment C5a des arg, or formyl-methionyl-leucyl-phenylalanine (FMLP), in concentrations ranging from 10(-10) to 10(-6) M, increased neutrophil adherence to endothelium in a dose-dependent manner. Purified C5a and C5a des arg were essentially equal in their ability to enhance neutrophil adherence, in contrast to the previously described greater in vitro potency of C5a compared with C5a des arg in stimulating neutrophil chemotaxis and enzyme release. Nonstimulated neutrophils adhered preferentially to human endothelial cells compared with fibroblasts or smooth muscle cells, suggesting that endothelial cells may make a unique contribution to the base-line adhesive interaction. However, chemotactic factors appear to enhance neutrophil adherence to endothelium by exerting an effect primarily on the neutrophil. In the presence of chemotactic factor, neutrophils adhered equally well to different cell types or to protein-coated plastic. Pretreatment of endothelial cells with chemotactic factor for as long as 4 h failed to increase subsequent neutrophil adherence. In contrast, pretreatment of neutrophils with chemotactic factor increased adherence to endothelium. Chemotactic factor-stimulated neutrophil adherence to endothelium occurred rapidly (within 2 min), diminished upon removal of stimulus, but could be rapidly and maximally restimulated upon readdition of the original dose of chemotactic factor. Thus, adherence to endothelium stimulated by chemotactic factor would appear to be a dynamic neutrophil response capable of rapid modulation, possibly important to the ability of neutrophils to adhere to and then migrate through vessel walls to localize at sites of inflammation.
M G Tonnesen, L A Smedly, P M Henson