leukotriene-b4 and 11-hydroxy-5-8-12-14-eicosatetraenoic-acid

To investigate the cooperation of chondrocytes and polymorphonuclear cells (PMN) in the biosynthesis of leukotrienes (LT).. PMN, resting and interleukin-1beta-stimulated cultured human chondrocytes, and mixtures of both cell types were incubated with A23187 and/or 14C-arachidonic acid (14C-AA). To explore the presence of LTC4 synthase and LTA4 hydrolase, the chondrocytes were incubated with authentic LTA4. Eicosanoids were analyzed using high performance liquid chromatography techniques.. Chondrocytes formed only prostaglandin E2 and minor amounts of 15-HETE and 11-HETE, the production of all of which was inhibited by 1 microM indomethacin. Incubation of PMN and chondrocytes produced more LTC4 from endogenous and exogenous AA, and more LTB4 from endogenous AA, than incubation of PMN alone, which was consistent with the presence of LTC4 synthase and LTA4 hydrolase activities in chondrocytes. Chondrocytes also slightly increased the level of PMN production of all 5-lipoxygenase (5-LO)-derived products from endogenous AA.. Human chondrocytes form eicosanoids from AA only by the cyclooxygenase pathway. Chondrocytes cooperate in the transcellular biosynthesis of LT since they possess LTA4 hydrolase and LTC4 synthase activities and increase metabolism by the 5-LO pathway in PMN.

Topics: Arachidonic Acid; Calcimycin; Cartilage, Articular; Cells, Cultured; Chondrocytes; Chromatography, High Pressure Liquid; Culture Media, Conditioned; Dinoprostone; Epoxide Hydrolases; Glutathione Transferase; Humans; Hydroxyeicosatetraenoic Acids; Indomethacin; Knee Joint; Leukotriene B4; Leukotriene C4; Neutrophils

To evaluate the cellular and biochemical composition of bronchoalveolar fluid in high-altitude pulmonary edema (HAPE), we performed bronchoalveolar lavage in three climbers with HAPE in a research facility at 4400 m on Mount McKinley. Three healthy climbers were used as controls. The HAPE fluids contained marked increases in high-molecular-weight proteins, erythrocytes, and leukocytes, most of which were alveolar macrophages. The HAPE fluids also contained detectable amounts of leukotriene B4 and other lipoxygenase products of arachidonic acid metabolism, complement fragments (C5a), inhibitors of neutrophil chemotaxis, and acid proteases but not hydroxyproline, a constituent of collagen. The data from this study indicate that HAPE involves a transient "large pore" leak in the pulmonary circulation. Despite the presence of two potent mediators of inflammation, leukotriene B4 and C5a, HAPE is not characterized by the intense neutrophil accumulation that is typical of other forms of acute lung injury.

Topics: Adult; Altitude Sickness; Cell Count; Chemotactic Factors; Humans; Hydroxyeicosatetraenoic Acids; Hypoxia; Leukotriene B4; Lung; Male; Molecular Weight; Permeability; Proteins; Pulmonary Edema; Therapeutic Irrigation

Our previous data suggested that lipoxygenation of endogenously released arachidonic acid (AA) is a critical step in stimulus-secretion coupling in the pancreatic beta cell. In the current study using monolayer cultures of neonatal rat islet cells, exogenous arachidonic acid (AA) (5 micrograms/ml) potently stimulated insulin release in the presence of a substimulatory glucose concentration, and potentiated release induced by glucose. Since the latter stimulatory effect of AA is prevented by inhibitors of the lipoxygenase pathway, we examined the effects of various lipoxygenase pathway products on glucose-induced insulin secretion. The mediator was not one of the stable end-products of either limb of the lipoxygenase pathway: 12- or 5-hydroxyeicosatetraenoic acid (HETE) (0.5-2000 ng/ml) did not alter insulin release, whereas 11-HETE, 15-HETE, leukotriene (LT)B4 and the delta 6 trans isomers of LTB4, LTC4 and 11-trans LTC4 all inhibited insulin release. Furthermore, diethylcarbamazine, a selective leukotriene synthesis inhibitor, did not prevent AA- or glucose-induced insulin release, arguing against a role for LTs as the mediator of AA's stimulatory effect. However, the unstable intermediate 12-hydroperoxyeicosatetraenoic acid (12-HPETE), and positional isomers of 12-HPETE, potentiated glucose-induced insulin secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Arachidonic Acid; Arachidonic Acids; Cells, Cultured; Diethylcarbamazine; Glucose; Hydroxyeicosatetraenoic Acids; Insulin; Insulin Secretion; Islets of Langerhans; Leukotriene B4; Lipoxygenase; Rats

The effects of 5-, 8-, 9-, 11-, 12-, and 15-monohydroxyeicosatetraenoic acid (HETE) (0.1-100 nM) on mucous glycoprotein release from cultured human airways were determined. Each of the HETE was an active secretagogue of mucus at concentrations greater than 1-10 nM with 12- and 15-HETE, the most active. Both 5- and 9-hydroperoxyeicosatetraenoic acid (HPETE) were also active as secretagogues at 100 nM, although of somewhat lower potency. As cultured airways were capable of responding to HETE with mucous glycoprotein release, it was of interest to identify and quantitate airway HETE formation. Accordingly, airways were incubated with tracer quantities of [14C]arachidonate for 16-48 h, and the spontaneous formation of 5-, 12- and 11- and/or 15-HETE was measured by high-pressure liquid chromatography. Indeed, sizeable quantities of 11- and/or 15- greater than 5- greater than 12-HETE were generated. This HETE generation was increased by the addition of 25 micrograms/ml of arachidonate and was reduced somewhat after 18-21 d in continuous tissue culture. Reversed anaphylaxis of human airways using anti-human IgE markedly increased the HETE formation, resulting in the production of micromolar concentrations of 5- and 11- and/or 15-HETE. Thus, human airways not only are capable of responding to the presence of HETE with mucous glycoprotein release, but also generate (both spontaneously and in response to anaphylaxis) at least three species of HETE, and do so in quantities capable of acting as mucus secretagogues.

Topics: Anaphylaxis; Arachidonic Acids; Culture Techniques; Humans; Hydroxyeicosatetraenoic Acids; Immunoglobulin E; Leukotriene B4; Lung; Mucus

A procedure using high performance liquid chromatography (HPLC) is described for the separation of major primary cyclooxygenase metabolites (prostacyclin metabolite-6ketoPGF1 alpha, thromboxane B2, and prostaglandins F2 alpha, E2, and D2), leukotrienes (C4, B4, and D4), monohydroxyeicosatetraenoic acids (15-, 11-, 12-, and 5HETEs), and free arachidonic acid. It is therefore possible to quantitate major arachidonic acid metabolites by a single chromatographic procedure. Using this technique we have determined that a major arachidonic acid metabolite of human lung macrophages co-elutes with leukotriene B4.

Topics: Arachidonic Acids; Chromatography, High Pressure Liquid; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Lung; Macrophages; Prostaglandins; Prostaglandins F; SRS-A; Thromboxane B2

The following high performance liquid chromatography system was found suitable for separating most lipoxygenase metabolites of arachidonic acid: Techsphere 5-C18 column, eluting solvent methanol:water:acetic acid (65:35:0.06 v/v), pH 5.3. Comparisons with other packing materials and solvent systems are described. The method could be used to identify lipoxygenase products released from mouse macrophage cells stimulated with gamma-hexachlorocyclohexane. Detection limits between 1 and 10 ng were obtained.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Arachidonic Acid; Arachidonic Acids; Chromatography, High Pressure Liquid; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Macrophages; Mice; SRS-A

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Arachidonic Acids; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Models, Biological; Mucus

The arachidonic acid released from cellular phospholipids of specifically stimulated platelets and leukocytes is oxygenated enzymatically by two major pathways. A complex cycloxygenase converts some of the free arachidonic acid to labile endoperoxides that are transformed to prostaglandins, thromboxanes and prostacyclin (PGI2). Lipoxygenases convert part of the arachidonic acid to unstable hydroperoxy-eicosatetraenoic acids (OOHETEs) that are transformed to monohydroxyeicosatetraenoic acids (HETEs), oligohydroxy-eicosatetraenoic or -eicostatrienoic acids such as di-HETEs and tri-HETEs, and, in some instances, more complex humoral mediators, including slow-reacting substances. Both the nature of the HETEs and the ratio of the HETEs to the cyclo-oxygenase products are specific characteristics of each type of cell. In human neutrophils, the sum of the lipoxygenase products 5-HETE, 11-HETE and 5,12-di-HETE substantially exceeds the total amount of PGE2 and other cyclo-oxygenase metabolites that are generated concurrently, and the endogenous lipoxygenase products regulate neutrophil function. The present data indicate that vitamin E (alpha-tocopherol) bidirectionally modulates the activity of the lipoxygenase pathway of human neutrophils in vitro. Normal plasma concentrations of alpha-tocopherol enhance the lipoxygenation of arachidonic acid, whereas higher concentrations of alpha-tocopherol exert a suppressive effect that is consistent with its role as a hydroperoxide scavenger.

Topics: Arachidonic Acids; Cell Membrane; Chemotaxis, Leukocyte; Complement C5; Dose-Response Relationship, Drug; Humans; Hydroxyeicosatetraenoic Acids; Leukotriene B4; Lipoxygenase; Lipoxygenase Inhibitors; Neutrophils; Vitamin E