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Anterior pituitary cells were incubated in the presence of luteinizing hormone-releasing hormone and one of three inhibitors of arachidonic acid metabolism: indomethacin, an inhibitor of the cyclooxygenase system; nordihydroguaiaretic acid, an antioxidant that inhibits lipoxygenase; and icosatetraynoic acid, an acetylenic analogue of arachidonic acid that blocks all known pathways of arachidonic acid metabolism. Indomethacin was ineffective in blocking luteinizing hormone-releasing hormone-stimulated luteinizing hormone secretion. Nordihydroguaiaretic acid was only marginally capable of inhibiting luteinizing hormone-releasing hormone-stimulated luteinizing hormone secretion. Icosatetraynoic acid at 10 pM completely inhibited stimulated luteinizing hormone secretion. Addition of several epoxygenated arachidonic acid metabolites to cells in vitro resulted in secretion of luteinizing hormone equal to or greater than that induced by 10 nM luteinizing hormone-releasing hormone. The half-maximal effective dose for these compounds was approximately 50 nM. The 5,6-epoxyicosatrienoic acid was the most potent of the compounds tested. These studies suggest that luteinizing hormone-releasing hormone-stimulated luteinizing hormone release is closely coupled with the production of oxidized arachidonic acid metabolites. Moreover, one or more of the epoxygenated arachidonic acid metabolites might be a component of the cascade of reactions initiated by luteinizing hormone-releasing hormone that ultimately results in secretion of luteinizing hormone.Calcium ions, cyclic nucleotides, and combinations thereof are postulated to be the "second messenger" of luteinizing hormone-releasing hormone (LHRH) (1, 2). The action of LHRH in stimulating the release of luteinizing hormone (LH) from anterior pituitary gonadotrophs also appears to involve turnover of phosphatidylinositol (3) and release of arachidonic acid from membrane phospholipid stores (4). Previous studies have eliminated a role for prostaglandins in the action of LHRH because LHRH-stimulated LH secretion is insensitive to blockage of prostaglandin formation by indomethacin (5). At least two additional pathways of arachidonate metabolism are currently recognized: a lipoxygenase reaction leading to hydroperoxides and leukotrienes (6) and the recently reported NADPH-supported cytochrome P450-dependent epoxygenase pathway (7-9). Since arachidonic acid mobilization from cellular pools is apparently required for LHRH-stimulated LH secretion (4), we presumed that either arachidonic acid or one or more of its metabolites of the lipoxygenase or epoxygenase pathways were active in stimulated secretion of LH. The studies reported here suggest that conversion of arachidonic acid to physiologically active metabolites constitutes an essential link in the cascade of intracellular events leading to LHRH-stimulated LH release. 99.9% by gas chromatographic analysis) was added in some cultures to a final concentration of 1 ,uM. The synthetic epoxyicosatrienoic acids (EETs) were pre...

Cancer cells need cholesterol to make new membrane. They get it either by de novo synthesis or from low-density lipoprotein (LDL), or both. Some types of cancer have very high LDL requirements. LDL particles, which circulate in the blood, contain a cholesteryl ester core surrounded by a phospholipid coat containing apoproteins that are recognized by LDL receptors on cell surfaces. After attachment to cells, LDL is endocytosed into lysosomes, where the core is exposed and hydrolyzed. A technique is known whereby LDL can be isolated, its core removed and replaced by a compatible lipophilic substance, and then reconstituted into intact LDL particles that are recognized and internalized by cells in the normal manner. A series of cytotoxic compounds has been synthesized, designed to be compatible with reconstituted LDL, and directed against cancers that copiously internalize LDL. They were evaluated by measuring the toxicity of reconstituted LDL toward test cells bearing LDL receptors. Selectivity was determined by comparison, either with mutant cells with few LDL receptors or with reconstituted methylated LDL (which is not recognized by LDL receptors) on normal cells. Two compounds, 19 and 25, were found that reconstitute well, kill or arrest the test cells at reasonably low concentrations, and are completely selective, suggesting that they are delivered to cells exclusively via the LDL pathway.

Various fatty acids were fed to the yeast Dipodascopsis uninucleata UOFS Y 128, and the extracted samples were analyzed for the accumulation of 3-hydroxy metabolites with the help of electron impact gas chromatography-mass spectrometry. Fatty acids containing of 5Z,8Z-diene system (5Z,8Z,11Z-eicosatrienoic, 5Z,8Z,11Z,14Z-eicosatetraenoic, and 5Z,8Z,11Z,14Z,17Z-eicosapentaenoic acids) yielded the corresponding 3-hydroxy-all-Z-eicosapolyenoic acids. Moreover, linoleic acid (9Z,12Z-octadecadienoic acid) and 11Z,14Z,17Z-eicosatrienoic acid were converted to the 3-hydorxylated metabolites of shorter chain length, e,g., 3-hydroxy-5Z,8Z-tetradecadienoic acid and 3-hydroxy-5Z,8Z,11Z-tetradecatrienoic acid, respectively. In contrast, no accumulation of a 3-hydroxy metabolite was observed with oleic acid (9Z-octadecenoic acid), linolelaidic acid (9E,12E-octadecadienoic acid), gamma-linolenic acid (6Z,9Z,12Z-octadecatrienoic acid), and eicosanoic acid as substrate. These findings pinpoint that the 3-hydroxylation of a fatty acid in Dipodascopsis uninucleata requires a 5Z,8Z-diene system either directly or following initial incomplete beta-oxidation. Following analysis of the enantiomer composition, the arachidonic acid metabolite was identified as 3R-hydroxy-5Z,8Z,11Z,14Z-eicosatetraenoic acid, which rules out a normal beta-oxidation as biosynthetic route to this new class of oxylipins.

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The authors have recently provided evidence implicating the cytochrome P-450 system in the generation of contractile tension of the ductus arteriosus. To confirm this possibility, carbon monoxide (CO/O2 ratio, 0.27) and cytochrome P-450 inhibitors [4-phenylimidazole; 14-isocyano,15-(methoxymethyleneoxy)-5Z,8Z,11 Z-eicosatrienoic acid; 9-hydroxyellipticine; alpha-naphthoflavone] were tested on the isolated ductus arteriosus from mature fetal lambs equilibrated at low (4-26 mm Hg) or high (229-694 mm Hg) O2 partial pressure (PO2). Carbon monoxide completely relaxed intact vessel wall preparations and preparations consisting of only the muscle. Carbon monoxide relaxation was reversed by illumination with monochromatic light and the peak for the photoactivated contraction occurred at 450 nm. 4-Phenylimidazole (100 and 1,000 microM) was also a relaxant agent, and its action was manifest at both low and high PO2. Unlike 4-phenylimidazole, the isonitrile compound (5 microM) and 9-hydroxyellipticine (10 and 100 microM) were relaxant only at low PO2 and were also less potent. At the same PO2, alpha-naphthoflavone (10 microM) barely reduced ductal tension. Treatment of the ductus with either a combination of superoxide dismutase (60 or 150 U/ml) and catalase (40 or 1,000 U/ml) or mannitol alone (80 mM) failed to alter the steady-state tone at low PO2 and the contractile response to O2. Arachidonic acid was tested on tissues pretreated with the dual cyclooxygenase and lipoxygenase inhibitor, BW755C (10 microM), and produced a weak relaxation at a concentration of 1 microM or higher. 5,6-Epoxytrienoic acid relaxed the untreated tissue, and its action was abolished by indomethacin (2.8 microM).(ABSTRACT TRUNCATED AT 250 WORDS)