Cloricromene, a coumarin derivative with antiaggregating and vasodilating properties, was tested in vitro on polymorphonuclear cell (PMN) adhesion to the endothelium, superoxide anion generation and chemotaxis. PMN adhesion was measured using cultured human umbilical vein endothelial cells (EC) either untreated or previously activated with interleukin-1 (IL-1). Cloricromene (5-50 microM) induced dose-related inhibition of PMN adhesion to untreated and IL-1 treated EC. Cloricromene also inhibited PMN superoxide generation induced by the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or by N-formyl-methionyl-leucyl-phenylalanine (FMLP). PMN and monocyte chemotaxis was evaluated by a modification of the Boyden chamber technique. Cloricromene inhibited both types of cell motility induced by FMLP in a concentration-dependent fashion. The major cloricromene metabolite (cloricromene acid) had no effect on any of the biological parameters studied up to a concentration of 500 microM. HPLC measurement showed that cloricromene accumulated in PMN within a few minutes and levels of the drug were still high after 60 min. In contrast its acid metabolite was not taken up in a significant amount during incubation periods up to 60 min. We conclude that cloricromene inhibits a series of PMN activities in vitro. This effect might be of pharmacological interest in view of the role of PMN activation in different pathophysiological conditions.

lnterieukln-1 (IL-1) induced slow, lasting activation of human endothelial cells (EC) to release prostacyclin (PGI 2 ). This was accompanied by endogenous3 H-arachldonlc acid f H-AA) release and by a time-dependent Increase In the cells' ability to convert exogenous AA. The continuous presence of IL-1 was not required, but about a 1-hour stimulation with the cytoklne was sufficient to trigger the cells to synthesize PGI 2 for several hours. The spectrum of 3 H-AA conversion shows that, In addition to 6-ketoprostaglandln F 1a , prostaglandln F^, also was raised after IL-1. The recovery of PGI 2 synthesis after aspirin was faster In IL-1-treated EC than In control cells. These data define some of the characteristics of IL-1 stimulation of PGI 2 and suggest that this process Is mediated both by endogenous AA mobilization and by an increase In cyclooxygenase activity. (Arteriosclerosis 10:129-134, January/February 1990) P rostacyclin (PGy is an important biosynthetic product of endothelial cells (EC). It inhibits platelet aggregation and thrombus formation and acts as a potent vasodilator by causing relaxation in vascular smooth muscle cells.1 PGI 2 hatf-life, when released into the circulation, is in the range of a few minutes, and its activity on platelets and vascular tone is extremely short lived. 1 Therefore, its biological relevance in vivo is strictly related to "how much and for how long" it can be produced by the vessel wall.Most known activators, e.g., thrombin, bradykinin, or ionophore A23187, induce a very fast, short-lasting release of PGI 2 by EC, followed by cell refractoriness to subsequent stimulation.2 ' 3 We and other groups have shown that the immunomediator, interieukin-1 (IL-1), has a peculiar way of inducing PGI 2 synthesis in EC.45 IL-1 stimulation requires a few hours to become apparent, but then it lasts for several hours.IL-1 is probably one of the most important mediators of the hemodynamic and hematological changes characteristic of septic shock.6 ' 7 The hemodynamic alterations, particularly the hypotension caused by IL-1, require cyclooxygenase products, since they are reversed by inhibitors of this enzyme.7 It has, therefore, been hypothesized that PGI 2 synthesis by EC plays an important role in mediating the slow and lasting drop in systemic vascular resistance that follows IL-1 release in the circulation. Despite its biological importance, little is known about the mechanism and the pattern through which IL-1 exerts its stimulatory activity on EC. In this work, we report that IL-1 induces endogenous arachidonic add (AA) mobilization and increases its conversion to prostaglandins via an increase in cyclooxygenase synthesis and activity. This effect is long lasting but does not need the continuous presence of IL-1 to be apparent. Methods Endothelial Cell CultureEC were isolated from human umbilical vein and cultured in medium 199 supplemented with 20% newborn calf serum as previously described in detail.2 The cells were used at the first passage at confluence, and they were maintaine...

Leukocytes and vascular cells interact closely in inflammation and immunity and lymphokines are important mediators of this interaction. The present study was designed to define the possible role of IL-6 as a communication signal between vascular and immunocompetent cells. IL-6 was measured as hybridoma growth factor (HGF) on the 7TD1 cell line in the supernatants of human endothelial cells (HEC). HEC released appreciable levels of HGF activity in the absence of deliberate stimulation. In vitro exposure to recombinant IL-1 beta markedly increased (usually 10 to 15-fold) HGF production by HEC. Optimal stimulation was observed with 0.1 to 50 U/ml for 4 to 20 h of incubation. Human and murine rIL-1 alpha stimulated HGF production in HEC. Anti-IL-6 antibodies inhibited the HGF activity of the HEC supernatants, thus confirming, together with the cytokine specificity of the assay, the nature of HEC-produced cytokine. IL-1-treated HEC expressed high levels of IL-6 mRNA as detected by Northern blot analysis. Inasmuch as IL-1 elicits a complex series of changes in HEC, it was important to assess whether IL-6, produced after exposure to IL-1, modified HEC function. Natural or rIL-6 did not affect the functional status of HEC as assessed by proliferative capacity, production of procoagulant activity and prostacyclin, ability to induce adhesion of polymorphonuclear leukocytes. The capacity to produce IL-6 may represent an important mechanism by which endothelial cells participate in inflammatory and immune reactions.