Human umbilical vein endothelial cells cultured on a collagen lattice were used to study the effects of the interaction between human monocytes and endothelial cells on the production of type 1 plasminogen activator inhibitor (PAI-1) by endothelial cells. The effects of adherence and transendothelial migration of monocytes on endothelial PAI-1 release were compared with those of other leukocytes, conditioned media from monocytes, and interleukin-1 beta (IL-1 beta). Because the cell culture system used allows simultaneous analysis of the lumenal and the subendothelial compartment of endothelial cell monolayers, we also studied into which direction PAI-1 is released by endothelial cells. Under quiescent conditions, the net amount of PAI-1 accumulated at the lumenal side was twofold higher than that accumulated at the subendothelial side (about 2.0 micrograms PAI-1/10(6) cells and 1.1 microgram PAI-1/10(6) cells, respectively, in 24 hours), as analyzed by a quantitative immunoradiometric assay (IRMA). Direct cell-cell contact between highly purified monocytes and endothelial cells strongly enhanced the PAI-1 release by endothelial cells in a dose-dependent way, whereas lymphocytes and neutrophils did not affect endothelial PAI- 1 production. The monocyte-mediated increase was first detected after 12 hours of incubation and lasted for at least 48 hours. In the presence of two monocytes per endothelial cell, the increases of PAI-1 at the lumenal side and at the subendothelial side were 87% and 32% in 24 hours, respectively. The effect of IL-1 beta on PAI-1 release by endothelial cells closely resembled that observed for monocytes. Monocyte-conditioned medium contained heat-labile product(s) which also, although to a much lesser extent than intact monocytes, enhanced endothelial PAI-1 release. Similarly, monocytes cultured on top endothelial cell separated by a microporous filter enhanced the release of PAI-1 to a lesser extent. Thus, these findings indicate that monocytes enhance endothelial PAI-1 release by mechanisms that are, at least in part, dependent on cell-cell contact.

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