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1.
Summary The precise role of eicosanoids in the development of myocardial injury during ischemia and reperfusion is still a matter of debate. Enhanced local production of these bioactive compounds appears to be a common response to tissue injury. Most likely, the cardiac tissue has the capacity to generate prostaglandins, thromboxanes as well as leukotrienes. Prostacyclin (PGI,) is the major eicosanoid produced by the jeopardized myocardium. In addition, at sites of tissue injury activation of platelets and infiltrating leukocytes results in the formation of considerable amounts of thromboxanes and leukotrienes. The production of eicosanoids requires prior release of arachidonic acid (AA) from phospholipids. Both ischemia and reperfusion are associated with a rise in the tissue level of AA. The absence of a proportional relationship between the tissue level of AA and the amounts of PGI, produced suggests that the sites of AA accumulation and PGI2 formation are different. It is conceivable that AA accumulation is mainly confined to myocytes, whereas the capacity to synthesize PGI, mainly resides in vascular cells. Both beneficial and detrimental effects of eicosanoids on cardiac tissue have been described. Prostaglandins act as vasodilators. Besides, some of the prostaglandins, especially PGI,, are thought to possess cyto-protective properties. Thromboxanes and leukotrienes may impede blood supply by increasing smooth muscle tone. Besides, leukotrienes augment vascular permeability. Experimental studies, designed to evaluate the effect of pharmacological agents, like PGI2-analogues and lipoxygenase and cyclo-oxygenase inhibitors, indicat that eicosanoids influence the outcome of myocardial injury. However, the delineation of the physiological significance of the locally produced eicosanoids is complicated by such factors as the wide variety of AA derivatives produced and the dose-dependency of their effects.  相似文献   

2.
Prostaglandins, thromboxanes (TX) and leukotrienes, collectively referred to as eicosanoids, are cyclooxygenase (COX) metabolites of arachidonic acid (AA). Prostaglandins, have been recognised for many years as key molecules in regulating reproductive tract physiology and pathology. Numerous recent studies in in vitro model systems and knockout mouse models have demonstrated specific functional roles for the respective cyclooxygenase enzymes, prostaglandins and prostanoid receptors. Here we review the findings obtained in several of these studies with emphasis on the roles played by cyclooxygenase enzymes and prostaglandins, specifically prostaglandin E2 (PGE2) and F2alpha in reproductive tract physiology and pathology.  相似文献   

3.
Arachidonic acid metabolism produces several biologically important compounds including the leukotrienes and prostaglandins. Prostaglandin H2 (PGH2) is the first metabolite in the arachidonic acid cascade leading to all other prostaglandins. Pivotal to our understanding of PGH2's biology is the ability to separate it in pure form from the numerous other arachidonic acid metabolites produced in a biological milieu. The extensive literature on PGH2 biology and metabolism has relied almost exclusively on the traditional method of separation using gravity flow silicic acid columns. In our hands, such PGH2 preparations were found to contain varying amounts of 12-hydroxy-5,8,10-heptadecatrienoic acid (HHT), PGE2, PGF2 alpha and other minor impurities as determined by further chromatographic and mass spectral analyses. Analytical separation of PGH2 and other arachidonic acid metabolites has been accomplished using reversed-phase HPLC. However, the labile nature of this molecule in aqueous systems makes such techniques unacceptable for preparative isolation of high purity PGH2 and has necessitated the development of a totally nonaqueous separation. To this end, we attempted several stationary phases and found that the cyano-bonded phase showed the best selectivity for resolving PGH2 from its major contaminants. Separations were performed on self-packed columns using a hexane-isopropanol gradient. Peaks were detected both by liquid scintillation counting and uv spectrophotometry (214 nm). Structure assignments were made by chromatographic comparison with authentic standards (PGF2 alpha, PGE2), biological activity (PGH2--platelet aggregation), and by ammonia direct chemical ionization mass spectrometry (HHT, hydroxy-5,8,10,14-eicosatetraenoic acid, PGH2, PGE2, PGF2 alpha). The latter technique, which by its very nature volatilizes all organic material in the sample, was particularly useful in determining not only that the PGH2 preparations were free from the aforementioned side products, but that they were also free from lipid, protein, and other potential residues frequently found in biological preparations.  相似文献   

4.
《ImmunoMethods》1993,2(3):187-194
Eicosanoids, both prostaglandins and leukotrienes, have been implicated as mediators in a number of physiological processes. Various tissues have been found to produce different types and quantities of eicosanoids. Tissues also differ in their eicosanoid profiles when the eicosanoids are produced under different conditions. The total amount of prostaglandins formed in response to cellular stimuli depends upon the release of arachidonic acid and its metabolism by PGH synthase (cyclooxygenase). Currently available assays for PGH synthase activity are too expensive, cumbersome, and insensitive to be used in screening a large number of samples for enzyme activity. The most sensitive assays available for prostaglandin detection are radioimmunoassays for specific prostaglandins. We discuss in this article the development of radioimmunoassays using monoclonal antibodies, both specific and pan-specific, for recognition of the prostaglandins.  相似文献   

5.
Prostaglandin is a term applied to a series of compounds derived enzymatically and nonenzymatically from 20 carbon fatty acids such as arachidonic acid (20:4). Research over the past 20 years has identified the prostaglandins as products of almost every cell although the amount and class of prostaglandin produced varies considerably with cell type. Within the last ten years additional, related products of 20:4 metabolism (operationally termed eicosanoids) have been discovered, eg. prostacyclin, thromboxanes, malondialdehyde, leukotrienes, hydroperoxy and hydroxy fatty acids. The role of these 20:4 metabolites in physiological and pathological states is currently under intensive investigation and the biological action of these compounds have been implicated in many key regulatory processes. Therefore, it is not unreasonable to predict that these potent bioregulatory compounds may have a central role in the initiation and regulation (positive and negative) of the spectrum of diseases which we functionally term “cancer”.  相似文献   

6.
Eicosanoids are a family of oxygenated metabolites of arachidonic acid, including the prostaglandins, thromboxanes, leukotrienes and lipoxins. These lipid mediators play essential roles in normal cellular homeostasis as well as in a number of disease states. This review will focus on recent advances in the field of eicosanoids and highlight specific discoveries and achievements. Emphasis will be placed on structure and receptor biology, which are of significant pharmacological and clinical relevance.  相似文献   

7.
Eicosanoids, including the prostaglandins, leukotrienes, hydroxyeicosatetraenoic acids, epoxyeicosatetraenoic acids, and related compounds, are biosynthetic, bioactive mediators derived from arachidonic acid (AA), a 20:4(n-6) fatty acid. We have developed a comprehensive and sensitive mass spectral analysis to survey eicosanoid release from endotoxin-stimulated RAW 264.7 macrophage-like cells that is capable of detecting over 70 diverse eicosanoids and eicosanoid metabolites, should they be present. We now address the question: Are biologically significant eicosanoids being overlooked? Herein, we illustrate a general approach to diverse isotope metabolic profiling of labeled exogenous substrates using mass spectrometry (DIMPLES/MS), demonstrated for one substrate (AA) and its resultant products (eicosanoids). RAW cells were incubated in medium supplemented with deuterium-labeled AA. When the cells are stimulated, two sets of eicosanoids are produced, one from endogenous AA and the other from the supplemented (exogenous) deuterium-labeled form. This produces a signature mass spectral "doublet" pattern, allowing for a comprehensive and diverse eicosanoid search requiring no previous knowledge or assumptions as to what these species may be, in contrast to traditional methods. We report herein observing unexpected AA metabolites generated by the cells, some of which may constitute novel bioactive eicosanoids or eicosanoid inactivation metabolites, as well as demonstrating differing metabolic pathways for the generation of isomeric prostaglandins and potential peroxisome proliferator-activated receptor activators. Unexpectedly, we report observing a series of 1a, 1b-dihomologue prostaglandins, products of adrenic acid (22:4(n-6)), resulting from the two-carbon elongation of AA by the RAW cells.  相似文献   

8.
The present studies were undertaken to examine the hypothesis that ethanol could affect cellular biosynthesis in the murine mastocytoma cell of prostaglandins and leukotrienes, oxidative metabolites of arachidonic acid, at concentrations that could be encountered in vivo as well as during in vitro experiments. The effects of ethanol which encompass these concentration ranges (200-1000 mg%) can be summarized as follows: first in the absence of exogenous arachidonic acid, ethanol caused a dose dependent decrease in the production of leukotrienes which was statistically significant at 200 mg%. At 1000 mg%, ethanol caused a 20-50% decrease in leukotrienes and a 21% decrease in the amount of prostaglandin D2 (PGD2) formed in these cells. Secondly, when cells were incubated with exogenous arachidonic acid (14 micrograms/ml), large increases in both PGD2 and leukotrienes occurred. Under these conditions, ethanol caused a further increase in the amount of leukotrienes and a small increase in the amount of PGD2 formed. This stimulatory effect was specific for ethanol since neither t-butanol nor n-butanol caused the enhanced production of leukotrienes with exogenous arachidonic acid. Thus, these experiments suggest that ethanol affects metabolism of arachidonic acid at reasonably low doses (200-400 mg%) of ethanol in a manner dependent on the free arachidonic acid in the tissue. Also, in vitro experiments in which ethanol is used as a solvent for arachidonic acid could be greatly affected by high levels of ethanol (500-1000 mg%) which are frequently utilized.  相似文献   

9.
The metabolism of arachidonic acid was studied using basal and differentiated keratinocytes as well as sebaceous cells isolated from hairless mice. These disassociated cells metabolized arachidonic acid predominantly to the prostaglandin H synthase products prostaglandins E2 and D2. 12-Hydroxyheptadecatrienoic acid (HHT), prostaglandin F2 alpha, thromboxane B2 and 6-ketoprostaglandin F1 alpha were also detected. Smaller amounts of the lipoxygenase products 5-, 12- and 15-hydroxyeicosatetraenoic acids (HETEs) were also detected. The major lipoxygenase product observed was 12-HETE. No leukotrienes or dihydroxy fatty acids were observed. The identity of the metabolites was established using several high-pressure liquid chromatography solvent systems. The biosynthesis of prostaglandins E2 and D2 was very rapid and was inhibited by the addition of indomethacin to the cells. The mixed population of keratinocytes and sebaceous cells were separated into enriched fractions by metrizamide gradients and elutriation techniques. The small, undifferentiated cells had high prostaglandin H synthase and 12-lipoxygenase activity. The basal cell-enriched fractions had the highest activity. With increasing differentiation of the cells, decreased biosynthetic activity was observed. These results indicate that undifferentiated keratinocytes, that is, the basal cells, may be an important source of prostaglandins and 12-HETE but are not a source of leukotrienes for the hairless mouse. It also suggests a role for keratinocyte-derived eicosanoids in the normal physiology of epidermal differentiation.  相似文献   

10.
Mouse macrophages pre-labeled with [3H]arachidonic acid (20:4) were shown to release metabolites generated by the lipoxygenase and cyclo-oxygenase pathways following in vitro addition of heat-killed Salmonella typhi. These metabolites were maximally released after 60-90 min of incubation and consisted of prostaglandins (85%), leukotriene C (6%), di-HETEs, leukotrienes D and E (4%), mono-HETEs (2%) and other metabolites (3%). Of the metabolites generated by the cyclo-oxygenase pathway (prostaglandins), 6-keto PGF1 alpha and PGE2 were generated at a ratio of 1.2 to 1. The significance and importance of these results are discussed.  相似文献   

11.
Considerable progress has been made in characterizing the individual participant enzymes and their relative contributions in the generation of eicosanoids, lipid mediators derived from arachidonic acid, such as prostaglandins and leukotrienes. However, the role of individual phospholipase (PL) A(2) enzymes in providing arachidonic acid to the downstream enzymes for eicosanoid generation in biologic processes has not been fully elucidated. In this review, we will provide an overview of the classification of the families of PLA(2) enzymes, their putative mechanisms of action, and their role(s) in eicosanoid generation and inflammation.  相似文献   

12.
Resident mouse peritoneal macrophages when exposed to zymosan during the first day of cell culture synthesize and secrete large amounts of prostaglandin E2 (PGE2) and leukotriene C4 (LTC4), the respective products of cyclo-oxygenase- and 5-lipoxygenase-catalysed oxygenations of arachidonic acid. Under these conditions of cell stimulation only small amounts of hydroxyeicosatetraenoic acids (HETEs) are concomitantly produced. However, exogenously added arachidonic acid is metabolized to large amounts of 12- and 15-HETE and only relatively small amounts of PGE2. No LTC4 is formed under these conditions. In contrast, resident mouse peritoneal macrophages in cell culture for 4 days synthesized less PGE2 and LTC4 when exposed to zymosan. However, these macrophage populations continue to synthesize 12-HETE from exogenously added arachidonic acid. Zymosan induced the secretion of a lysosomal enzyme, N-acetyl-beta-glucosaminidase, equally in both 1- and 4-day cultures. Both 12- and 15-hydroperoxyeicosatetraenoic acids (HPETEs), the precursors of 12- and 15-HETE, were found to be irreversible inhibitors of the cyclo-oxygenase pathway and reversible inhibitors of the 5-lipoxygenase pathway in macrophages. 15-HETE were found to be reversible inhibitors of both pathways. Thus the oxidation of arachidonic oxidation of arachidonic acid to both prostaglandins and leukotrienes may be under intracellular regulation by products of 12- and 15-lipoxygenases.  相似文献   

13.
The release of arachidonic acid, a precursor in the production of prostaglandins and leukotrienes, is achieved by activity of the cytosolic phospholipase A(2)α (cPLA(2)α). Signaling mediated by this class of bioactive lipids, which are collectively referred to as eicosanoids, has numerous effects in physiological and pathological processes. Herein, we report the development of a ligand-based pharmacophore model and pharmacophore-based virtual screening of the National Cancer Institute (NCI) database, leading to the identification of 4-(hexadecyloxy)-3-(2-(hydroxyimino)-3-oxobutanamido)benzoic acid (NSC 119957) as cPLA(2)α inhibitor in cell-free and cell-based in vitro assays.  相似文献   

14.
The present studies were undertaken to examine the hypothesis that ethanol could effect cellular biosynthesis in the murine mastocytoma cell of prostaglandins and leukotrienes, oxidative metabolites of arachidonic acid, at concentrations that could be encountered in vivo as well as during in vitro experiments. The effects of ethanol which encompass these concentration ranges (200–1000 mg%) can be summarized as follows: first in the absence of exogenous arachidonic acid, ethanol caused a dose dependent decrease in the production of leukotrienes which was statistically significant at 200 mg%. At 1000 mg%, ethanol caused a 20–50% decrease in leukotrienes and a 21% decrease in the amount of prostaglandins D2 (PGD2) formed in these cells. Secondly, when cells were incubated with exogenous arachidonic acid (14 μg/ml), large increases in both PGD2 and leukotrienes occurred. Under these conditions, ethanol caused a further increase in the amount of leukotrienes and a small increase in the amount of PGD2 formed. This stimulatory effect was specific for ethanol since neither t-butanol nor n-butanol caused the enhanced production of leukotrienes with exogenous arachidonic acid. Thus, these experiments sugsests that ethanol affects metabolsim of arachidonic acid at reasonably low doses (200–400 mg%) of ethanol in a manner dependent on the free arachidonic acid in the tissue. Also, in vitro experiments in which ethanol is used as a solvent for arachidonic acid could be greatly affected by high levels of ethanol (500–1000 mg%) which are frequently utilized.  相似文献   

15.
Enhanced penetration by ethanol of exogenous arachidonic acid into human leukocyte preparations results in the production of large amounts of eicosanoids including 5-, 12- and 15-hydroxyeicosatetraenoic acids as well as the leukotrienes C4 delta 6-trans-leukotriene B4, 12-epi-delta 6-trans-leukotriene B4, leukotriene B4 and 5(S), 12(S)-dihydroxyeicosatetraenoic acid. The production of these compounds is affected by the concentrations of both ethanol and arachidonic acid independently in a complex manner with stimulation at lower concentrations and later relative inhibition. It was shown that the resulting leukotriene B4 exhibited the same specific activity as exogenous arachidonic acid when labelled substrate was used.  相似文献   

16.
17.
Inflammatory lipid mediators derived from arachidonic acid (AA) and docosahexaenoic acid (DHA) modify the pathophysiology of brain ischemia. The goal of this work was to investigate the formation of eicosanoids and docosanoids generated from AA and DHA, respectively, during no-flow cerebral ischemia. Rats were subjected to head-focused microwave irradiation 5 min following decapitation (complete ischemia) or prior to decapitation (controls). Brain lipids were extracted and analyzed by reverse-phase liquid chromatography-tandem mass spectrometry. After complete ischemia, brain AA, DHA, and docosapentaenoic acid concentrations increased 18-, 5- and 4-fold compared with controls, respectively. Prostaglandin E(2) (PGE(2)) and PGD(2) could not be detected in control microwaved rat brain, suggesting little endogenous PGE(2)/D(2) production in the brain in the absence of experimental manipulation. Concentrations of thromboxane B(2), E(2)/D(2)-isoprostanes, 5-hydroxyeicosatetraenoic acid (5-HETE), 5-oxo-eicosatetraenoic acid, and 12-HETE were significantly elevated in ischemic brains. In addition, DHA products such as mono-, di- and trihydroxy-DHA were detected in control and ischemic brains. Monohydroxy-DHA, identified as 17-hydroxy-DHA and thought to be the immediate precursor of neuroprotectin D(1), was 6.5-fold higher in ischemic than in control brain. The present study demonstrated increased formation of eicosanoids, E(2)/D(2)-IsoPs, and docosanoids following cerebral ischemia. A balance of these lipid mediators may mediate immediate events of ischemic injury and recovery.  相似文献   

18.
Prostaglandins, leukotrienes and mono-hydroxy acid products of arachidonic acid were measured in mucus of freshly recovered morning stools of a patient with an exacerbation of ulcerative proctocolitis. Eicosanoids in ether extracts were separated by high performance liquid chromatography and amounts determined by radioimmunoassay. Four hydroxy-eicosatetraenoic acids were detected, of which the most important one was identified as 15-hydroxy eicosatetraenoic acid (530 ng/g mucus). Leukotriene B4 was also present (21 ng/g mucus) and small amounts of immunoreactive leukotriene C4 (less than 0.8 ng/g mucus). The prostaglandins 6-keto-PGF1 alpha and PGE2 and thromboxane B2 were found in amounts of 3.7, 2.0 and 9.2 ng/g mucus, respectively.  相似文献   

19.
The data on whether T cells produce leukotrienes or other 5-lipoxygenase metabolites of arachidonic acid is conflicting. We report that exogenous arachidonic acid added to phytohemagglutin-stimulated human T cells profoundly inhibits leukotriene B4 production, with 90% inhibition caused by 10(-6) M arachidonic acid. The 12- and 15-lipoxygenase pathways were also inhibited by arachidonic acid. Recent reports that human T cells produce no 5-lipoxygenase metabolites of arachidonic acid might be explained by the fact that the studies used greater than or equal to 10(-5)M arachidonic acid in the incubation media.  相似文献   

20.
The metabolic pathway called the arachidonic acid cascade produces a wide range of eicosanoids, such as prostaglandins, thromboxanes and leukotrienes with potent biological activities. Recombinant DNA techniques have made it possible to determine the nucleotide sequences of cDNAs and/or genomic structures for the enzymes involved in the pathway. Sequence comparison analyses of the accumulated sequence data have brought great insights into the structure, function and molecular evolution of the enzymes. This paper reviews the sequence comparison analyses of the enzymes involved in the arachidonic acid cascade.  相似文献   

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