Spontaneous and lipopolysaccharide-induced expression of procoagulant activity by equine lung macrophages in comparison with blood monocytes and blood neutrophils

Abstract

The procoagulant activity (PCA) associated with equine bronchoalveolar lavage cells was determined and compared with that expressed by peripheral blood mononuclear cells and neutrophils. Lung cell preparations from horses affected with chronic pulmonary disease were included in all experiments and there was no difference in the qualitative type of response compared with lung cells which were obtained from healthy horses. Significant amounts of PCA were expressed by cells freshly procured from bronchoalveolar lavages of healthy and diseased horses. When adherent lung cells were kept in culture for some time, cell-associated PCA slightly decreased within 4 h, reached its lowest point after approximately 24 h and rose again during the second week of culture. In contrast, freshly isolated blood mononuclear cells or neutrophils expressed little PCA. Following culture for 24 h, mononuclear cells began to express increased PCA levels. Both cultivated lung cells (comprised mainly on alveolar macrophages) and blood mononuclear cells responded to LPS by dramatically increased PCA expression, whereas neutrophils showed a small augmentation of PCA on LPS stimulation. Fresh mononuclear cells and cultivated lung cells differed in their PCA response to LPS in several respects. Blood mononuclear cells were more sensitive to LPS than lung macrophages and responded to a 100-fold lower LPS concentration than the latter. Mononuclear cell-associated PCA peaked 4 h after stimulation whereas that of cultured macrophages continued to increase up to 24 h after stimulation. Lung macrophages cultured in adherence responded to LPS stimulation with a much higher PCA increase than macrophages cultured in suspension, in teflon containers. However, the culture vessel did not influence the PCA expressed by unstimulated cells. PCA expression depended to a large extent on transcription and translation, as evidenced by a 60-85% reduction of PCA in cycloheximide- or actinomycin D-treated, LPS-stimulated lung macrophages. PCA was largely cell-associated; only a small proportion of cell-associated PCA was shed into the medium. The PCA associated with mononuclear cells and with lung macrophages was tissue factor because of its dependence on clotting factor VII and its independence from clotting factor VIII. The expression of PCA by freshly isolated cells, the lower sensitivity to LPS, and the loss of PCA in the first 24 h of cultivation are indicative of in vivo activation of lung macrophages.