Purification, characterization and selective inhibition of human prostaglandin G/H synthase 1 and 2 expressed in the baculovirus system

Barnett, Jim; Chow, Joan M.; Ives, D. A. J.; Chiou, Melody; Mackenzie, Rebecca B.; Osen, Eric; Nguyen, Binh; Tsing, Stan; Bach, Chinh; Freire, José Ednésio da Cruz; Chan, Hardy; Sigal, Elliott; Ramesha, Chakkodabylu S.

doi:10.1016/0167-4838(94)90148-1

Cited by 315 publications

(230 citation statements)

References 32 publications

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“…1995 . Our data also indicate that, although breaking the integrity of the cell plasma membrane should facilitate the access of metamizol to the endoplasmic reticulum and nuclear envelope, where cyclooxygenase-2 is located Ž . Morita et al, 1995 , cell purified enzymes have also been described for other Ž NSAIDs Barnett et al, 1994;Mitchell et al, 1994;. Carabaza et al, 1996 .…”

Section: Discussionmentioning

confidence: 99%

Regulation of cyclooxygenase activity by metamizol

Campos

Gregorio

Garcı́a-Nieto

et al. 1999

European Journal of Pharmacology

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The ability of metamizol to inhibit cyclooxygenase-1 and cyclooxygenase-2 activities has been evaluated using different cyclooxygenase sources. Metamizol inhibited purified cyclooxygenase-1 and cyclooxygenase-2 with an IC of about 150 mgrml. A similar IC 50 50value for cyclooxygenase-2 was obtained in lipopolysaccharide-activated broken murine macrophages. Consistent with these findings, molecular models of the complexes between cyclooxygenase-1 or cyclooxygenase-2 with 4-methylaminoantipyrine, the major active derivative of metamizol, suggested a common binding mode to both isoforms. In intact cells, however, the inhibition profiles were Ž . markedly different. The IC values of metamizol for cyclooxygenase-1 in intact bovine aortic endothelial cells BAEC cells and human 50 platelets were 1730 " 150 mgrml and 486 " 56 mgrml, respectively. Inhibition of cyclooxygenase-2 activity in murine macrophages and primary human leukocytes activated by lipopolysaccharide yielded IC values of 12 " 1.8 mgrml and 21 " 2.9 mgrml, 50 respectively. These data indicate that the IC values obtained with purified enzymes or disrupted cells cannot always be extrapolated to 50 Ž . the cyclooxygenase inhibitory activity of nonsteroidal antiinflammatory drugs NSAIDs in intact cells. The data presented here also indicate that cyclooxygenase-2 inhibition could play an important role in the pharmacological effects of metamizol. q 1999 Elsevier Science B.V. All rights reserved.Ž .

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Section: Discussionmentioning

confidence: 99%

Regulation of cyclooxygenase activity by metamizol

Campos

Gregorio

Garcı́a-Nieto

et al. 1999

European Journal of Pharmacology

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“…Furthermore, it partially prevented the enhancement of myelopoiesis normally observed in adiponectin-containing cultures (Table II). In addition to Dup-697, the SC-58125 and NS-398 COX-2 selective inhibitors (42,43) as well as the APHS dual COX-1/COX-2 inhibitor (44) restored B lymphopoiesis in adiponectin-contained cultures (Table II). Interestingly, SC-560, a selective inhibitor for COX-1 (IC 50 ϭ 9 nM; COX-2, IC 50 ϭ 6.3 M), (45) also abrogated adiponectin activity at 10 Ϫ7 M (Table II).…”

Section: A Potential Role For Pgs In Adiponectin-mediated Responsesmentioning

confidence: 99%

Adiponectin, a Fat Cell Product, Influences the Earliest Lymphocyte Precursors in Bone Marrow Cultures by Activation of the Cyclooxygenase-Prostaglandin Pathway in Stromal Cells

Yokota

Meka

Kouro

et al. 2003

The Journal of Immunology

130

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Adiponectin, an adipocyte-derived hormone, is attracting considerable interest as a potential drug for diabetes and obesity. Originally cloned from human s.c. fat, the protein is also found in bone marrow fat cells and has an inhibitory effect on adipocyte differentiation. The aim of the present study is to explore possible influences on lymphohematopoiesis. Recombinant adiponectin strongly inhibited B lymphopoiesis in long-term bone marrow cultures, but only when stromal cells were present and only when cultures were initiated with the earliest category of lymphocyte precursors. Cyclooxygenase inhibitors abrogated the response of early lymphoid progenitors to adiponectin in stromal cell-containing cultures. Furthermore, PGE2, a major product of cyclooxygenase-2 activity, had a direct inhibitory influence on purified hematopoietic cells, suggesting a possible mechanism of adiponectin action in culture. In contrast to lymphopoiesis, myelopoiesis was slightly enhanced in adiponectin-treated bone marrow cultures, and even when cultures were initiated with single lymphomyeloid progenitors. Finally, human B lymphopoiesis was also sensitive to adiponectin in stromal cell cocultures. These results suggest that adiponectin can negatively and selectively influence lymphopoiesis through induction of PG synthesis. They also indicate ways that adipocytes in bone marrow can contribute to regulation of blood cell formation.

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“…Recombinant COX-1 and COX-2 Enzymes-Recombinant human COX-1 and COX-2 enzymes were expressed in a baculovirus system and partially purified as described previously (17). The specific activity of the enzymes used were between 13,000 and 21,000 units (1 unit ϭ 1 nmol of oxygen consumed/mg of protein/min) with a purity of Ϸ60% as judged by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining.…”

Section: Methodsmentioning

confidence: 99%

“…Enzymatic Assays-The purified COX enzymes were reconstituted with 2 mM phenol and 1 M hematin (Sigma), and the cyclooxygenase activity was measured either by the oxygen uptake assay or by the radiometric assay exactly as described previously (17). K m values were determined radiometrically using 2-200 M 1-14 C-labeled AA or AEA (specific activity, 3-4 ϫ 10 5 cpm/nmol).…”

Section: Methodsmentioning

confidence: 99%

Synthesis of Prostaglandin E2 Ethanolamide from Anandamide by Cyclooxygenase-2

Ives

Ramesha

1997

Journal of Biological Chemistry

382

277

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Because of its structural similarity to polyunsaturated fatty acids, anandamide could serve as substrate for enzymes such as lipoxygenases and cyclooxygenases, which metabolize polyunsaturated fatty acids to potent bioactive metabolites. Here the ability of recombinant human cyclooxygenase-1 (hCOX-1) and cyclooxygenase-2 (hCOX-2) to metabolize anandamide was studied. Baculovirus-expressed and -purified hCOX-2, but not hCOX-1, effectively oxygenated anandamide. Reverse phase high pressure liquid chromatography analysis of the products derived from 1-14 C-labeled anandamide showed that the products formed are similar to those formed with arachidonic acid as substrate. The major prostanoid product derived from anandamide was determined by mass spectrometry to be prostaglandin E 2 ethanolamide. Incubation of anandamide with lysates and the intact cell line expressing COX-2 but not that of COX-1 produced prostaglandin E 2 ethanolamide. These results demonstrate the existence of a COX-2-mediated pathway for anandamide metabolism, and the metabolites formed represent a novel class of prostaglandins.Anandamide (arachidonoyl ethanolamide, AEA) 1 is a polyunsaturated fatty acyl amide that was identified from porcine brain lipids as an endogenous ligand for brain cannabinoid receptor (1). Although structurally different from cannabinoids, AEA by its ability to activate the central CB1 receptor displays pharmacological properties similar to cannabinoids (2, 3). In addition to its central action via the CB1 receptor, AEA displays potent immunomodulatory and anti-inflammatory activities by interacting with peripheral CB1 and/or CB2 receptors (4 -6).Free AEA is present in both central and peripheral tissues (see Ref. 7 for a review) and could interact with CB receptors to display some of its immunomodulatory and anti-inflammatory activities. In addition, AEA is also stored esterified to phosphatidylethanolamines and is released by the action of phospholipase D in response to various stimuli (7). The AEA thus released inside the cell could participate in signal transduction as a second messenger and display some of its immunomodulatory and anti-inflammatory activities independent of its interaction with the CB receptors. In fact, AEA has been shown to antagonize CB2 receptors, and it is not clear how this antagonism results in immunomodulatory activities observed in cells only expressing CB2 receptors (8). It is possible that a metabolite of AEA rather than AEA itself could account for all or some of these properties. Furthermore, because of its structural similarities to polyunsaturated fatty acids, endogenously released AEA could serve as substrate for lipoxygenases and cyclooxygenases (COX) that metabolize polyunsaturated fatty acids to potent bioactive molecules. It has been demonstrated that lipoxygenase could metabolize AEA, and the metabolites have potent biological activities (9, 10). However, it is not known whether COX can metabolize AEA. Arachidonic acid (AA) is the substrate for both COX-1 and COX-2. AEA is structur...

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Purification, characterization and selective inhibition of human prostaglandin G/H synthase 1 and 2 expressed in the baculovirus system

Cited by 315 publications

References 32 publications

Regulation of cyclooxygenase activity by metamizol

Regulation of cyclooxygenase activity by metamizol

Adiponectin, a Fat Cell Product, Influences the Earliest Lymphocyte Precursors in Bone Marrow Cultures by Activation of the Cyclooxygenase-Prostaglandin Pathway in Stromal Cells

Synthesis of Prostaglandin E2 Ethanolamide from Anandamide by Cyclooxygenase-2

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