Biomedical subjects

R F Myers

Publications and source records attributed to R F Myers.

17 recordsLinked to original sources

PubMed1986

Metabolism of platelet-activating factor (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) by human promyelocytic leukemic HL60 cells. Stimulated expression of phospholipase A2 and acetyltransferase requires differentiation.

Human promyelocytic leukemia (HL60) cells can be induced to differentiate into mature granulocytes by exposure to dimethyl sulfoxide. The addition of N-formylMet-Leu-Phe or the Ca2+ ionophore A23187 to these differentiated cells generated 15-30 pmol of platelet-activating factor (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine (alkylacetyl-GPC)/10(6) cells as quantified by platelet aggregation assays. Under identical conditions, uninduced cells produced little alkylacetyl-GPC. Upon the addition of ionophore A23187, differentiated cells, and not uninduced ones, released [14C]arachidonate from prelabeled phospholipids including ether-linked phosphatidylcholines, formed both 3H-labeled 1-O-alkyl-2-lyso-sn-glycero-3-phosphocholine (alkyllyso-GPC) and [3H]alkylacetyl-GPC from endogenous 3H-labeled 1-O-alkyl-2-(long chain) acyl-sn-glycero-3-phosphocholine (alkylacyl-GPC), and incorporated exogenously added [3H]acetate or [3H]alkyllyso-GPC into alkylacetyl-GPC. These results are suggestive that both phospholipase A2 and acetyltransferase activities are involved in alkylacetyl-GPC biosynthesis by HL60 cells and that these activities appear during differentiation. However, when measured in cell extracts, the activities of phospholipase A2 and acetyltransferase of uninduced cells were virtually indistinguishable from those of differentiated cells. Uninduced cells exhibited enhanced incorporation of [3H]alkyllyso-GPC or [3H]alkylacetyl-GPC into alkylacyl-GPC and of [14C]arachidonate and [14C]oleate into various phospholipids including phosphatidylcholine. However, such enhanced expression of acylation reactions could not account for the lack of accumulation of arachidonate or of alkylacetyl-GPC by uninduced cells. Furthermore, analyses of phospholipid classes by phosphorus determination showed no significant alterations in phospholipid composition of HL60 cells during differentiation. Together these data are suggestive that mechanisms regulating the activation of phospholipase A2 and acetyltransferase activities are defective in uninduced cells and that an increased concentration of cytosolic free Ca2+ alone is not a sufficient requirement for these mechanisms.

Acetyltransferases↗

PubMed1985

Ex vivo effects of nonsteroidal antiinflammatory drugs on arachidonic acid metabolism in neutrophils from a reverse passive Arthus reaction.

Rat neutrophils isolated from 4-h reverse passive Arthus reaction (RPAR) pleural exudates actively metabolize arachidonic acid via cyclooxygenase and lipoxygenase. Utilizing this system, the effect of oral doses of nonsteroidal antiinflammatory drugs on the ability of these cells to produce HHT, 5-HETE, and LTB from exogenously added arachidonic acid has been investigated. In vitro and ex vivo, indomethacin and timegadine inhibit cyclooxygenase activity in rat pleural neutrophils. In vitro, timegadine is a lipoxygenase as well as a cyclooxygenase inhibitor. This dual inhibition is confirmed by the observation that ex vivo timegadine inhibits the production of lipoxygenase as well as cyclooxygenase metabolites. While indomethacin, a cyclooxygenase inhibitor, primarily inhibits edema formation, the inhibition of both pathways of arachidonic acid metabolism by timegadine is reflected in the drug's ability to reduce cellular influx as well as edema formation in the RPAR pleural cavity inflammatory reaction.

Animals↗

PubMed1985

Ionophore-stimulated lipoxygenase activity and histamine release in a cloned murine mast cell, MC9.

A cloned murine mast cell line designated MC9 expresses a 5-lipoxygenase activity when stimulated with the ionophore A23187. Upon addition of 0.5 microM ionophore, MC9 cells produce 270 +/- 43 pmoles 5-HETE, 74 +/- 40 pmoles 5,12 diHETEs and 65 +/- 31 pmoles LTC4/10(6) cells from 37 microM exogenously added [1-14C]arachidonic acid in two minutes. 5-HETE and 5,12-diHETES, including LTB4 were identified by GC/MS whereas LTC4 was confirmed by HPLC mobility, bio-assay, RIA and enzymatic transformation. The principal cyclooxygenase products were PGD2 and TxB2 (8.5 +/- 2.4 and 5.4 +/- 1.2 pmoles/10(6) cells respectively). Prostanoids were identified by comigration with authentic standards on two-dimensional thin layer chromatograms. Production of arachidonic acid lipoxygenase metabolites stimulated with ionophore proved relatively insensitive to removal of extracellular Ca+2 and chelation by EGTA. In addition, MC9 5-lipoxygenase required only low micromolar amounts of exogenous arachidonic acid for maximal activity. Whereas production of arachidonic acid metabolites lasted only two to five minutes, histamine release stimulated with ionophore was not initiated until 5 minutes (12 +/- 3% cellular histamine) and continued for 30 minutes (37 +/- 7% cellular histamine). Although these cells metabolize arachidonic acid differently from the classic peritoneal-derived mast cell, they resemble subpopulations found in certain tissues (such as mucosa) and should be useful in understanding the biochemistry of mast cell mediator release.

Animals↗

PubMed1984

The appearance of phospholipase activity in the human macrophage-like cell line U937 during dimethyl sulfoxide induced differentiation.

The human histiocyte cell line, U937, with monocyte characteristics, can be induced to differentiate into macrophage-like cells when exposed to growth medium containing 1.5% DMSO. Following three days of exposure, DMSO-treated but not control U937 cells can be stimulated to release endogenous arachidonic acid from their phospholipids. Maximum release of the unsaturated fatty acid occurs with 10 microM calcium ionophore in the presence but not in the absence of exogenously added calcium ion. In addition, DMSO-treated but not control U937 cells exhibit phospholipase activity when exposed to human IgG and then anti-human immunoglobulin. These data suggest that with respect to arachidonic acid metabolism U937 cells differentiate into functional macrophage-like cells when exposed to DMSO.

Arachidonic Acid↗

PubMed1983

Differential effects of anti-inflammatory drugs on lipoxygenase and cyclo-oxygenase activities of neutrophils from a reverse passive Arthus reaction.

Rat neutrophils isolated from four-hour reverse passive Arthus reaction pleural exudates actively metabolize arachidonic acid. Production of 11-hydroxy- and 15-hydroxy-icosatetraenoic acid and 12-hydroxy-heptadecatrienoic acid is inhibited by indomethacin, benoxaprofen, BW 755C, piroxicam, ibuprofen, timegadine, and naproxen, suggesting that production of these arachidonic acid metabolites occurs at similar enzymic active sites. In addition, in the presence of the calcium inophore A23187 or the non-ionic detergent, BRIJ 56, rat neutrophils also produce the lipoxygenase products 5-hydroxy-icosatetraenoic acid and leukotriene B. The production of these metabolites is calcium dependent. Moreover, the calcium ionophore A23187 and BRIJ 56 synergistically act to augment the metabolism of exogenously added arachidonic acid via lipoxygenase. The formation of these metabolites is inhibited by BW 755C, benoxaprofen and timegadine but not by other non-steroidal anti-inflammatory drugs tested. In fact, at doses which inhibit cyclo-oxygenase activity, indomethacin, naproxen, and ibuprofen stimulate arachidonic acid metabolism via lipoxygenase.

Animals↗

PubMed1981