INDUCTION OF PROSTAGLANDIN ENDOPEROXIDE SYNTHASE-2 IN HUMAN MONOCYTESASSOCIATED WITH CYCLO-OXYGENASE-DEPENDENT F-2-ISOPROSTANE FORMATION

1 The isoprostane 8-epi-prostaglandin (PG)F-2 alpha is produced by fre e radical-catalyzed peroxidation of arachidonic acid. It may also be f ormed as a minor product of the cyclo-oxygenase activity of platelet P GH synthase (PGHS)-1. We investigated 8-epi-PGF(2 alpha) production as sociated with induction of the human monocyte PGHS-2 and its pharmacol ogical modulation. 2 Heparinized whole blood samples were drawn from h ealthy volunteers, 48 h following oral dosing with aspirin 300 mg to s uppress platelet cyclo-oxygenase activity. One mi aliquots were incuba ted with lipopolysaccharide (LPS: 0.1 - 50 mu g ml(-1)) for 0 - 24 h a t 37 degrees C. PGE(2) and 8-epi-PGF(2 alpha) were measured in separat ed plasma by radioimmunoassay and enzyme immunoassay techniques. 3 Lev els of both eicosanoids were undetectable (i.e. < 60 pg ml(-1)) at tim e 0. LPS induced the formation of PGE(2) and 8-epi-PGF(2 alpha) in a t ime- and concentration-dependent fashion, coincident with the inductio n of PGHS-2 detected by Western blot analysis of monocyte lysates. Aft er 24 h at 10 mu g ml(-1) LPS, immunoreactive PGE(2) and 8-epi-PGF(2 a lpha) averaged 10,480 +/- 4,643 and 295 +/- 140 pg ml(-1) (mean +/- s. d., n = 6), respectively. 4 Dexamethasone and sulphonamido-6-(2,4-difl uorothiophenyl)-1-indanone (L-745,337), a selective inhibitor of the c yclo-oxygenase activity of PGHS-2, reduced PGE(2) and 8-epi-PGF(2 alph a) production in response to LPS. 5 Isolated monocytes produced PGE(2) and 8-epi-PGF(2 alpha) in response to LPS (10 pg ml(-1)) in a time-de pendent fashion. Monocyte PGE(2) and 8-epi-PGF(2 alpha) production was largely prevented by dexamethasone (2 mu M) and cycloheximide (10 mu g ml(-1)) in association with suppression of PGHS-2 but not of PGHS-1 expression. 6 We conclude that the induction of PGHS-2 in human monocy tes is associated with cyclo-oxygenase-dependent generation of the vas oconstrictor and platelet-agonist 8-epi-PGF(2 alpha).