Importance of L-selectin–dependent leukocyte–leukocyte interactions in human whole blood

The objective of this study was to investigate whether leukocytes could be recruited by rolling leukocytes in a human whole blood model system. In all experiments, either neutrophils, whole blood, or diluted blood was perfused over immobilized E-selectin. With isolated neutrophils (2 x 10(5)/mL), the free-flowing neutrophils were captured by attached neutrophils to form secondary interactions that resulted in lines of rolling leukocytes. These secondary tethers accounted for 50% to 60% of all interactions and were eliminated by an L-selectin antibody, which also eliminated the lines of rolling leukocytes. Perfusion of whole blood or diluted blood revealed no lines of rolling leukocytes. The addition of red blood cells to isolated neutrophils either in a 1000:1 or a 10:1 ratio also inhibited lines of rolling leukocytes. Leukocytes were fluorescently labeled with rhodamine-6G so that leukocyte-leukocyte interactions could be studied in whole blood. A small number of secondary tethers (less than 20%) occurred and could be reduced by more than 80% with an L-selectin antibody. However, the overall impact on leukocyte recruitment was negligible. Similar experiments were performed using murine whole blood or isolated murine leukocytes. In the absence of red blood cells, murine leukocytes also formed lines of rolling leukocytes on E-selectin, and secondary tethers accounted for 50% of total interactions. However, when murine blood (diluted 1:5 with buffer) was perfused over E-selectin, secondary tethers accounted for only 13% of total interactions. These interactions were completely absent when blood was used from L-selectin-deficient mice. These data demonstrate for the first time that the importance of L-selectin-dependent leukocyte-leukocyte interactions is greatly reduced in whole blood and does not enhance overall recruitment of leukocytes in this physiologic milieu. (Blood. 2000;95:2954-2959)