Generation of the isoprostane 8-epi-prostaglandin F2alpha in vitro and in vivo via the cyclooxygenases.
F2-isoprostanes are isomers of the prostaglandin PGF2alpha. At least one compound of this group, 8-epi-PGF2alpha, exhibits biological activity, and therefore special interest is focused on the mechanism of isoprostane formation: enzyme catalyzed or radical mediated. We analyzed the formation of isoprostanes in vitro and in vivo. In both systems, purified cyclooxygenase isoenzymes and cell models specific for the cyclooxygenase isoenzymes, 8-epi-PGF2alpha formation could be totally suppressed by cyclooxygenase inhibitors. Indomethacin inhibited concentration-dependent 8-epi-PGF2alpha formation in platelets stimulated with calcium ionophore, arachidonic acid or thrombin. Nordihydroguaiaretic acid, an antioxidant, blocked isoprostane formation with a similar IC50 value as thromboxane B2 synthesis, pointing toward cyclooxygenase as the primary target of inhibition. Based on the turnover number, cyclooxygenase-2 formed higher levels of 8-epi-PGF2alpha than cyclooxygenase-1. Endogenous 8-epi-PGF2alpha production in rat mesangial cells correlated well with the mRNA and protein expression of cyclooxygenase-2 during interleukin-1 induction. However, in contrast to human platelets, which produced different forms of isoprostanes, rat mesangial cells appeared to form only 8-epi-PGF2alpha. Further, this indicates that mesangial cells may represent a cellular origin for renal 8-epi-PGF2alpha formation. Next, we analyzed the formation of isoprostanes in humans. A direct correlation was observed between indomethacin treatment and the decrease in 8-epi-PGF2alpha and isoprostane levels, but compared with other prostanoids the inhibition was less pronounced. In summary, based on the in vitro studies, a clear cyclooxygenase-dependent formation of isoprostanes, especially 8-epi-PGF2alpha, was observed. However, in vivo additional formation via cyclooxygenase enzyme-independent mechanisms is likely.
Klein
T
T
Department of Pediatrics, Philipps University of Marburg, Germany.
Reutter
F
F
Schweer
H
H
Seyberth
H W
HW
Nüsing
R M
RM
D000293:Adolescent; D000328:Adult; D000818:Animals; D001792:Blood Platelets / Q000378:metabolism; D002648:Child; D015237:Dinoprost / Q000031:analogs & derivatives* / Q000096:biosynthesis; D005920:Glomerular Mesangium / Q000378:metabolism; D006801:Humans; D007213:Indomethacin / Q000494:pharmacology; D007527:Isoenzymes / Q000502:physiology; D008297:Male; D011451:Prostaglandin-Endoperoxide Synthases / Q000502:physiology*; D051381:Rats
D007527:Isoenzymes; C075750:8-epi-prostaglandin F2alpha; D015237:Dinoprost; D011451:Prostaglandin-Endoperoxide Synthases; D007213:Indomethacin