6-ketoprostaglandin-f1-alpha and 11-hydroxy-5-8-12-14-eicosatetraenoic-acid

Arachidonic acid metabolism via cyclooxygenase, lipoxygenase, and cytochrome P-450 epoxygenase was investigated in thoracic aortic tissue obtained from rabbits fed either standard rabbit chow or chow containing 2% cholesterol. Aortic strips were incubated with [14C]arachidonic acid and A23187. Metabolites from extracted media were resolved by high-pressure liquid chromatography (HPLC). Normal and cholesterol-fed rabbit aortas synthesized prostaglandins (PGs) and hydroxyeicosatetraenoic acids (HETEs). The major cyclooxygenase products were 6-keto-PGF1 alpha and PGE2. Basal aortic 6-keto-PGF1 alpha production was slightly reduced in cholesterol-fed compared with normal rabbits. 12(S)- and 15(S)-HETE were the major aortic lipoxygenase products from both normal and cholesterol-fed rabbits. The structures were confirmed by gas chromatography-mass spectrometry (GC-MS). Only cholesterol-fed rabbit aortas metabolized arachidonic acid via cytochrome P-450 epoxygenase to the epoxyeicosatrienoic acids (EETs). 14,15-, 11,12-, 8,9-, and 5,6-EET were identified based on comigration on HPLC with known 14C-labeled standards and typical mass spectra. Incubation of normal aorta with 14,15-EET decreased the basal synthesis of 6-keto-PGF1 alpha. The other EETs were without effect. The four EET regioisomers relaxed the norepinephrine-precontracted normal and cholesterol-fed rabbit aorta. The relaxation response to 14,15-EET was greater in aortas from cholesterol-fed rabbits. These studies demonstrate that hypercholesterolemia, before the development of atherosclerosis, alters arachidonic acid metabolism via both the cyclooxygenase and epoxygenase pathways.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; 4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyrazol-3-amine; 6-Ketoprostaglandin F1 alpha; 8,11,14-Eicosatrienoic Acid; Animals; Aorta, Thoracic; Arachidonic Acids; Carbon Radioisotopes; Cholesterol, Dietary; Clotrimazole; Diet, Atherogenic; Hydroxyeicosatetraenoic Acids; In Vitro Techniques; Indomethacin; Kinetics; Masoprocol; Metyrapone; Muscle, Smooth, Vascular; Rabbits; Reference Values; Stereoisomerism

Pulp was experimentally inflamed by applying bacterial lipopolysaccharide (LPS). Changes in arachidonic acid (AA) metabolites were determined by measuring the conversion of exogenously added AA in pulp homogenates. The inflamed pulp produced 12-hydroxy-eicosatetraenoic acid (12-HETE), 6-keto-prostaglandin (PG) F1 alpha greater than PGE2, thromboxane B2 and 11-HETE, which was further identified with high-performance liquid chromatography. The LPS treatment caused a 2.0-fold increase in 12-HETE production at 1 h, a 3.8-fold increase in 6-keto-PGF1 alpha production at 12 h and increases in PGE2 and 11-HETE production of 8.8- and 5.5-fold, respectively, at 24 h. Vascular permeability in the inflamed pulp was measured by quantifying the amount of an extravasated dye; it increased markedly from 6 h and reached a peak at 12 h after the LPS application. When indomethacin (0.3-30 mg/kg, s.c.) was given before LPS, both the production of 6-keto-PGF1 alpha and PGE2 and the increase in vascular permeability were inhibited dose dependently. Exogenously applied PGE2 and PGI2 methyl ester reduced the inhibition of the increase in vascular permeability caused by indomethacin. Thus PGE2 and PGI2 may be involved in increases in vascular permeability in pulpal inflammation.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; 6-Ketoprostaglandin F1 alpha; Animals; Arachidonic Acid; Arachidonic Acids; Capillary Permeability; Chromatography, High Pressure Liquid; Dental Pulp; Dinoprostone; Dose-Response Relationship, Drug; Hydroxyeicosatetraenoic Acids; Indomethacin; Lipopolysaccharides; Male; Pulpitis; Rats; Rats, Inbred Strains; Thromboxane B2

Particulate fractions and slices from fetal calf aorta convert arachidonic acid to 6-oxoprostaglandin F1 alpha (6-oxoPGF1 alpha), 6,15-dioxoPGF1 alpha, 12-hydroxy-5,8,10-heptadecatrienoic acid, 11-hydroxy-5,8,12,14-icosatetraenoic acid (11h-20:4), and 15-hydroxy-5,8,11,13-icosatetraenoic acid (15h-20:4). In some cases, small amounts of 12-hydroxy-5,8,10,14-icosatetraenoic acid (12h-20:4) were also detected. The products were all identified by gas chromatography-mass spectrometry after purification by normal phase and argentation high pressure liquid chromatography. Both 11h-20:4 and 15h-20:4 appeared to be formed by prostaglandin endoperoxide synthetase rather than by lipoxygenases, since their formation was inhibited by indomethacin but not by nordihydroguaiaretic acid. The formation of 12h-20:4, on the other hand, was stimulated by indomethacin, probably due to increased substrate availability. The formation of hydroxyicosatetraenoic acids was markedly stimulated by adrenaline. Substantial amounts of 6,15-dioxoPGF1 alpha were formed from arachidonic acid by particulate fractions from fetal calf blood vessels, especially in the presence of relatively high substrate concentrations. The formation of this product was stimulated by methemoglobin and inhibited by adrenaline, glutathione, and tryptophan. It would appear that particulate fractions from fetal calf aorta convert arachidonic acid to 15-hydroperoxyPGI2, which can either be reduced in the presence of various cofactors to form PGI2 or dehydrated to give 15-oxoPGI2. The formation of hydroperoxides from arachidonic acid could be an important factor in regulating PGI2 synthesis in aorta, since PGI2 synthetase is strongly inhibited by such intermediates.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; 6-Ketoprostaglandin F1 alpha; Animals; Aorta; Arachidonic Acids; Cattle; Ductus Arteriosus; Fatty Acids, Unsaturated; Hydroxy Acids; Hydroxyeicosatetraenoic Acids; Kinetics; Lipoxygenase; Prostaglandin-Endoperoxide Synthases; Prostaglandins F, Synthetic