Facilitation of granulocyte migration into bovine pulmonary artery intimal explants by intact viable endothelium

To characterize the role of normal endothelium in granulocyte chemotaxis, the authors measured granulocyte adherence to and migration into bovine pulmonary artery intimal explants. Explants were placed, endothelium uppermost, in chemotaxis chambers with zymosan-activated plasma in the lower well and 5 X 10(6)/ml 51Cr-labeled granulocytes in the upper well. After 15, 30, 60, 120, 180, or 240 minutes incubation at 37 C, granulocyte adherence was measured by removal of adherent granulocytes from the endothelial layer with a 0.1% trypsin wash and counting of radioactivity in the wash. Scanning electron microscopy confirmed that this technique removed the majority of adherent cells from the endothelial surface without disrupting its continuity. Migration was calculated by counting of the remaining radioactivity in the explant. Granulocyte migration with Medium 199 alone in the lower well (random migration) was 36 +/- 3% by 3 hours. Chemotaxis-induced migration at each time studied was 1.5-2 times random migration. Granulocyte adherence was between 4% and 9% in both groups at all times examined. In some experiments, either endothelium was removed from explants or explants were fixed with glutaraldehyde prior to experimentation. Removal of endothelium resulted in a two-fold increase in granulocyte adherence but no significant difference in migration, compared with intact intimal explants. Glutaraldehyde fixation of explants resulted in more tightly adherent granulocytes and significantly less migration. With lactate dehydrogenase as a marker of endothelial cell damage, granulocyte migration in response to zymosan-activated plasma did not injure endothelium. It is concluded that, in blood vessels, chemotaxis is an interactive process between granulocytes and endothelium and that intact, viable endothelium facilitates granulocyte migration.