Lymphocyte and granulocyte migration across the endothelial layer of bovine pulmonary artery intimal explants towards lymphocyte conditioned medium

Abstract

An intravenous infusion of endotoxin into sheep results in accumulation of equal numbers of lymphocytes and granulocytes in the pulmonary microcirculation. The role of the sequestered lymphocytes in acute lung injury is not known. The present study examines whether lymphocyte migration through pulmonary endothelium contributes to endothelial damage and also examines the effect of lymphokines on granulocyte migration. Bovine pulmonary artery intimai explants were mounted in Boyden chambers and conditioned media, prepared from bovine peripheral blood lymphocytes, was used as the chemoattractant. The rate of 51Cr labelled bovine granulocyte or lymphocyte migration into intimal explants was determined over a 3 hr incubation period. Permeability changes were assessed by adding trace amounts of 14C-sucrose and 3H-water to the upper well and following their rate of equilibration with the lower well. 6-Keto-PGF 1 a was measured in the upper well. Lymphocyte conditioned media was found to be chemotactic for both lymphocytes and granulocytes (lymphocyte migration at 60 min: lymphocyte conditioned media = 18.5 ± 2.3%, mean ± s.e. RPMI control = 12.5 ± 1.5; granulocyte migration at 120 min: conditioned media = 36.1 ± 5.7, RPMI control = 18.2 ± 3.0). Ultrastructural examination revealed leukocyte migration followed an orderly sequence during which the leukocytes maintained close contact with the adjacent endothelial cells. No structural evidence of endothelial cell damage was seen at any time examined. Granulocyte migration was associated with an increased rate of 14C-sucrose equilibration after 2 hr of incubation (lower well counts/upper well counts at 2 hr, RPMI control = 0.18 ± 0.02; lymphocyte conditioned medium = 0.30 ± 0.04) indicating alteration in the endothelial barrier function. Leukocyte migration, particularly lymphocyte migration, was accompanied by a marked increase in prostacyclin accumulation (3 hr: no leukocytes, 188 ± 17 ng/ml; lymphocytes, 560 ± 104). These in vitro findings suggest that lymphocytes and lymphokines may be involved in acute lung injury and also that permeability changes associated with granulocyte migration may depend on the chemoattractant.