Induction of prostaglandin endoperoxide synthase-2 in human monocytes associated with cyclo-oxygenase-dependent F2-isoprostane formation.
1. The isoprostane 8-epi-prostaglandin (PG)F2 alpha is produced by free radical-catalyzed peroxidation of arachidonic acid. It may also be formed as a minor product of the cyclo-oxygenase activity of platelet PGH synthase (PGHS)-1. We investigated 8-epi-PGF2 alpha production associated with induction of the human monocyte PGHS-2 and its pharmacological modulation. 2. Heparinized whole blood samples were drawn from healthy volunteers, 48 h following oral dosing with aspirin 300 mg to suppress platelet cyclo-oxygenase activity. One ml aliquots were incubated with lipopolysaccharide (LPS: 0.1-50 micrograms ml-1) for 0-24 h at 37 degrees C. PGE2 and 8-epi-PGF2 alpha were measured in separated plasma by radioimmunoassay and enzyme immunoassay techniques. 3. Levels of both eicosanoids were undetectable (i.e. < 60 pg ml-1) at time 0. LPS induced the formation of PGE2 and 8-epi-PGF2 alpha in a time- and concentration-dependent fashion, coincident with the induction of PGHS-2 detected by Western blot analysis of monocyte lysates. After 24 h at 10 micrograms ml-1 LPS, immunoreactive PGE2 and 8-epi-PGF2 alpha averaged 10,480 +/- 4,643 and 295 +/- 140 pg ml-1 (mean +/- s.d., n = 6), respectively. 4. Dexamethasone and 5-methanesulphonamido-6-(2,4-difluorothiophenyl)-1-indano ne (L-745,337), a selective inhibitor of the cyclo-oxygenase activity of PGHS-2, reduced PGE2 and 8-epi-PGF2 alpha production in response to LPS. 5. Isolated monocytes produced PGE2 and 8-epi-PGE2 alpha in response to LPS (10 micrograms ml-1) in a time-dependent fashion. Monocyte PGE2 and 8-epi-PGF2 alpha production was largely prevented by dexamethasone (2 microM) and cycloheximide (10 micrograms 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 monocytes is associated with cyclo-oxygenase-dependent generation of the vasoconstrictor and platelet-agonist 8-epi-PGF2 alpha.
Patrignani
P
P
Department of Pharmacology, University of Chieti G. D'Annunzio School of Medicine, Italy.
Santini
G
G
Panara
M R
MR
Sciulli
M G
MG
Greco
A
A
Rotondo
M T
MT
di Giamberardino
M
M
Maclouf
J
J
Ciabattoni
G
G
Patrono
C
C
D000328:Adult; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology; D015237:Dinoprost / Q000031:analogs & derivatives / Q000097:blood; D015232:Dinoprostone / Q000097:blood; D005260:Female; D006801:Humans; D007189:Indans / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008297:Male; D008875:Middle Aged; D009504:Neutrophils; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis*; D014662:Vasoconstrictor Agents
D016861:Cyclooxygenase Inhibitors; D007189:Indans; C093700:L 745337; D008070:Lipopolysaccharides; D014662:Vasoconstrictor Agents; C075750:8-epi-prostaglandin F2alpha; D003907:Dexamethasone; D015237:Dinoprost; D011451:Prostaglandin-Endoperoxide Synthases; D015232:Dinoprostone