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W S Powell

Publications and source records attributed to W S Powell.

At least 91 records · Page 5Linked to original sources
PubMed1984

Properties of leukotriene B4 20-hydroxylase from polymorphonuclear leukocytes.

Human polymorphonuclear leukocytes (PMNL) convert arachidonic acid (20:4) to a number of dihydroxy metabolites, including leukotriene B4 (LTB4) 5S,12R-dihydroxy-6,8,10,14-EEEZ-icosatetraenoic acid (isomer-1), 5S,12S-dihydroxy-6,8,10,14-EEEZ-icosatetraenoic acid, 5S,12S-dihydroxy-6,8,10,14-EZEZ-icosatetraenoic acid (5S,12S-dh-20:4), 5,6-dihydroxy-7,9,11,14-icosatetraenoic acid, and 5,15-dihydroxy-6,8,11,13-icosatetraenoic acid. LTB4 was synthesized rapidly after stimulation of PMNL with the divalent cation ionophore, A23187, but its concentration rapidly declined after about 4 min, in contrast to the other dihydroxy metabolites of 20:4 whose concentrations remained stable for at least 20 min. The amounts of polar metabolites (identified primarily as 20-hydroxy-LTB4) increased steadily with time up to 20 min. These results suggest that LTB4 may be specifically converted to its 20-hydroxy metabolite by PMNL. We prepared 3H- and 14C-labeled analogs of the dihydroxyicosatetraenoic acid metabolites described above by incubation of labeled 20:4 with PMNL. Although all of these substances were metabolized to some extent by human PMNL, LTB4 (apparent Km, 1.0 microM) was metabolized the most rapidly, followed by 5S,12S-dh-20:4 (apparent Km, 2.4 microM) and isomer-1 (apparent Km, 4.8 microM). All three substrates were shown by mass spectrometry to be converted to their 20-hydroxy metabolites. LTB4 was also metabolized to its omega-carboxy derivative. Human mononuclear leukocytes and rabbit PMNL metabolized LTB4 very slowly, whereas rat PMNL metabolized this substrate at about one-sixth the rate of human PMNL. These results demonstrate that human PMNL contain an omega-hydroxylase that specifically converts LTB4 to its 20-hydroxy metabolite. This enzyme may be important for the regulation of LTB4 levels in vivo.

Animals↗
PubMed1983

Metabolism of linoleic acid by prostaglandin endoperoxide synthase from adult and fetal blood vessels.

Linoleic acid (18:2) is converted by prostaglandin endoperoxide synthase in particulate fractions and homogenates of fetal calf aorta to its 9- and 13-hydroperoxy metabolites. These intermediates are then either dehydrated to the corresponding oxo compounds or reduced to monohydroxy products. Alternatively, the hydroperoxyoctadecadienoic acids can be converted to epoxyhydroxyoctadecenoic acids, which are hydrolyzed to trihydroxy metabolites by epoxide hydrolases present in both particulate and cytosolic fractions from aorta. Linoleic acid (Km, 442 microM) is a much poorer substrate for prostaglandin endoperoxide synthase than is arachidonic acid (20:4) (Km, 48 microM). However, the oxygenation of 18:2 by particulate fractions from aorta is linear with time for at least 5 min, whereas the oxygenation of 20:4 is linear for only 15 s. Arachidonic acid strongly inhibits the conversion of 18:2 to monohydroxy (ID50, 10 microM) and trihydroxy (ID50, 140 microM) products. Linoleic acid has a similar, but much weaker effect on the formation of 6-oxoprostaglandin F1 alpha from 20:4. Substantial amounts of both the monohydroxy (9-hydroxy-10, 12-octadecadienoic acid and 13-hydroxy-9,11-octadecadienoic acid) and trihydroxy (9,10,11-trihydroxy-12-octadecenoic acid, 9,10,13-trihydroxy-11-octadecenoic acid and 9,12,13-trihydroxy-10-octadecenoic acid) metabolites of 18:2 were shown by gas chromatography-mass spectrometry to be formed from endogenous substrate during incubation of slices of fetal calf aorta in physiological medium. This raises the possibility that some of these products or their hydroperoxy precursors may have some biological significance.

Aging↗
PubMed1983

Effects of protein deficiency on the metabolism of arachidonic acid by rat pleural polymorphonuclear leukocytes.

The effects of protein deficiency on the biosynthesis of metabolites of arachidonic acid by rat pleural polymorphonuclear leukocytes stimulated with calcium ionophore were investigated. The major products of metabolism by lipoxygenase in these cells were leukotriene B4 and 5-hydroxy-6,8,11,14-eicosatetraenoic acid, whereas the major cyclooxygenase products were thromboxane B2 and 12-hydroxy-5,8,10-heptadecatrienoic acid. At high substrate concentrations (100 microM), the formation of all products by polymorphonuclear leukocytes was lower for protein-deficient rats than for controls. Similar results were obtained when products synthesized from endogenous substrate were measured, except that there was no change in the amount of 5-hydroxy-6,8,11,14-eicosatetraenoic acid formed. The biosynthesis of prostaglandins E2 and F2 alpha by homogenates of rat kidney medulla was reduced as a result of protein deficiency. Acetylsalicylic acid inhibited the formation of cyclooxygenase products and stimulated the formation of lipoxygenase products by polymorphonuclear leukocytes. Protein deficiency did not alter the effects of acetylsalicylic acid on the biosynthesis of these products, although at any given concentration the amounts of products formed were less with protein-deficient rats than with rats fed control diets.

Animals↗
PubMed1983

Separation of unlabeled metabolites of arachidonic acid from their deuterium- and tritium-labeled analogs by argentation high-pressure liquid chromatography.

Deuterium- and tritium-labeled analogs of high isotopic purity are required for the analysis of metabolites of arachidonic acid (20:4) by mass spectrometry and radioimmunoassay. We prepared a number of 2H-labeled standards by incubating [5,6,8,9,11,12,14,15-2H]20:4 with soybean lipoxygenase, polymorphonuclear leukocytes, or homogenates of rat spleen or bovine lung. The unlabeled and 2H-labeled forms of the following products were completely resolved by argentation high-pressure liquid chromatography: 20:4; 5-hydroxy-6,8,11,14-eicosatetraenoic acid (5h-20:4); 11h-20:4; 12h-20:4; 15h-20:4; 12-hydroxy-5,8,10-heptadecatrienoic acid; leukotriene B4; prostaglandin D2 (PGD2); and PGF2 alpha. The corresponding forms of PGE2 and thromboxane B2 were also separated, but not quite as well as the above compounds. Analysis by mass spectrometry indicated that the amounts of unlabeled material in approx 1:1 mixtures of unlabeled and 2H-labeled products could be reduced to 0.1% or less after two steps of argentation high-performance liquid chromatography. Metabolites of 20:4 labeled with tritium were also separated from their unlabeled counterparts and had retention times longer than those of the corresponding 2H-labeled analogs.

Animals↗
PubMed1983

Conversion of 8,11,14-eicosatrienoic acid to 11,12-epoxy-10-hydroxy-8-heptadecenoic acid by aorta.

Particulate fractions from fetal calf aorta convert 8,11,14-eicosatrienoic acid to a number of products derived from 12-hydroperoxy-8,10-heptadecadienoic acid, including 2 stereoisomers of 11,12-epoxy-10-hydroxy-8-heptadecenoic acid (11,12e-10h-17:1), which were identified by mass spectrometry. In the early stages of the reaction, considerable amounts of the epoxyhydroxy isomers were formed, but the amounts of these products decreased as the reaction continued. There was a concomitant increase in the formation of 10,11,12-trihydroxy-8-heptadecenoic acid (10,11,12th-17:1), which was present only in small amounts initially. Incubation of the 2 isomers of 11,12e-10h-17:1 with microsomal and cytosolic fractions resulted in their conversion to isomers of 10,11,12th-17:1. No hydrolysis of the epoxides occurred in the presence of boiled tissue fractions.

8,11,14-Eicosatrienoic Acid↗
PubMed1983

Separation of unlabeled and isotopically-labeled metabolites of arachidonic acid by argentation high-pressure liquid chromatography.

Argentation HPLC can be used to separate unlabeled metabolites of 20:4 from their deuterium and tritium-labeled analogs. In many cases baseline resolution of labeled and unlabeled forms is achieved. This method can be used to obtain labeled compounds of high isotopic purity which are suitable for use as standards in radioimmunoassay or GC-MS using selected ion monitoring.

Animals↗
PubMed1982

Formation of 6-oxoprostaglandin F1 alpha, 6,15-dioxoprostaglandin F1 alpha, and monohydroxyicosatetraenoic acids from arachidonic acid by fetal calf aorta and ductus arteriosus.

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.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1981

Prostaglandin E-binding sites in the fetal adrenal.

Prostaglandin E2 (PGE2) is specifically and reversibly bound by particulate fractions from adrenals from fetal calves and lambs. These binding sites appear to be located primarily in the adrenal medulla and have properties very similar to those which we have detected in adult human, bovine, and ovine adrenal medulla. The rate constant for the association reaction between [3H]PGE2 and its binding sites in fetal calf adrenals is 1.8 X 10(5) M-1 sec-1. The [3H]PGE2-binding site complex dissociates in a biphasic manner, with rate constants of 6.7 X 10(-4) sec-1 and 4.2 X 10(-5) sec-1 for the fast and slow phases, respectively. The Kd values for the interactions of PGE2 with particulate fractions from adrenals of fetal calves and lambs are 2.3 and 3.5 nM, respectively. The [3H]PGE2-binding sites are very specific for PGE1 and PGE2. The affinities of other prostaglandins (PGF2 alpha greater than PGA2 greater than 13,14-dihydro-15-oxo-PGE2 greater than PGD2 greater than 6-oxo-PGF1 alpha) are 130- to 1300-fold lower. We have detected binding sites in adrenals from fetal calves as early as 112 days of gestation. The concentration of binding sites in fetal calf adrenals increases from 0.11 to 0.18 pmol/mg protein between about 130 days of gestation and term. The concentration of binding sites in fetal lamb adrenals (approximately 0.07 pmol/mg protein) does not change appreciably between about 100 days of gestation and term. These results suggest that PGE-binding sites are present in fetal adrenal medulla in mid- and late gestation and are consistent with a role for PGEs in controlling catecholamine release from the fetal adrenal.

Adrenal Cortex↗
PubMed1980

Rapid extraction of oxygenated metabolites of arachidonic acid from biological samples using octadecylsilyl silica.

A rapid procedure for the efficient extraction of prostaglandins, thromboxanes and hydroxy fatty acids from urine, plasma and tissue homogenates has been developed. Fractions containing these substances are acidified and passed through a column of octadecylsilyl silica, which retains oxygenated metabolites of arachidonic acid. Phospholipids, proteins and very polar materials either are not retained or can be eluted with dilute aqueous ethanol. Nonpolar lipids and monohydroxy fatty acids are then eluted with petroleum ether or benzene. Subsequent elution of the column with methyl formate gives a fraction containing prostaglandins and thromboxanes which is much less contaminated with extraneous material than that obtained by conventional extraction of aqueous media with organic solvents. The methyl formate can be removed rapidly under a stream of nitrogen and the components of the sample purified directly by high pressure liquid chromatography (HPLC). An improved method for the purification of prostaglandins and TXB2 by HPLC on silica columns is reported.

Animals↗
PubMed1980

Distribution of prostaglandin omega-hydroxylases in different tissues.

The conversion of prostaglandins E2 and F2 alpha to their 19- and 20-hydroxy metabolites by various tissues has been measured by gas chromatography-mass spectrometry (GC-MS) using selected ion monitoring. A number of different tissues of the pregnant rabbit possess prostaglandin 20-hydroxylase activity (lung > liver > fetal placenta > maternal placenta approximately equal to uterus > renal cortex > renal medulla approximately equal to placental membranes). With the exception of the liver, prostaglandins E2 and F2 alpha are metabolized at equal rates by the 20-hydroxylases of different tissues. Only lung and liver microsomes posess high levels of prostaglandin 20-hydroxylase in non-pregnant rabbits and only liver microsomes have appreciable 19-hydroxylase activity. Pulmonary prostaglandin 20-hydroxylase is induced in male rabbits by treatment with progesterone. On the basis of substrate specificity studies and the effects of a cytochrome P-450 inhibitor, SKF-525A, the prostaglandin 20-hydroxylases of lung and liver microsomes from pregnant rabbits appear to be different enzymes. In pregnant rats and hamsters, liver and kidney are the only tissues in which we detected prostaglandin omega-hydroxylase activity.

Animals↗
PubMed1980

Effects of prostaglandins on the adenylate cyclase activity of lungs from fetal rabbits.

Prostaglandin E1 (PGE1) and PGI2 stimulate adenylate cyclase activity in lungs from fetal rabbits (29 days of gestation) with EC50 values of 4.4 and 2.6 X 10(-7) M, respectively. PGE2, PGD2, PGF2 alpha, and 6-oxoPGF1 alpha are active only at much higher concentrations. The response of adenylate cyclase to PGE1 rises gradually from 16% to 60% above basal between days 21-26 of gestation. Between days 26-27 of gestation, there is a sharp increase in stimulated activity to 120% above basal (P < 0.001). This level is maintained until 3 days after birth. A further increase was observed 8 days after birth. Stimulation of adenylate cyclase by fluoride increases gradually from about 100% to nearly 400% above basal activity between 21 days of gestation and 8 days after birth. During this time there are no significant changes in basal activity. The increased response of adenylate cyclase to PGE1 between days 26-27 of gestation occurs at a time when unbound plasma cortisol and synthesis of pulmonary surfactant start to increase in the fetal rabbit. It is possible that cortisol sensitizes adenylate cyclase to prostaglandins, which could explain some of the its actions on surfactant synthesis.

Adenylyl Cyclases↗