PubMed Health⌕ Search

Biomedical subjects

S Fiore

Publications and source records attributed to S Fiore.

16 recordsLinked to original sources
PubMed1992

Lipoxin recognition sites. Specific binding of labeled lipoxin A4 with human neutrophils.

Lipoxin A4 stimulates rapid lipid remodeling and a pertussis toxin-sensitive release of arachidonic acid in polymorphonuclear leukocytes (PMN) (Nigam, S., Fiore, S., Luscinskas, F.W., and Serhan, C.N. (1990) J. Cell. Physiol. 143, 512-523) and has been shown to inhibit leukocyte responses in several systems. To examine the basis underlying these actions, we have prepared [11,12-3H]lipoxin A4 (LXA4) and characterized its interactions with human PMN. Time course studies (0-90 min) with intact PMN demonstrated cell association of 3H label which was specific and reversible. PMN bound [3H]LXA4 with a Kd of 0.5 +/- 0.3 nM, representing approximately 1,830 sites/PMN, and the Hill plot value of 1.9 suggests cooperative binding. [3H]LXA4 binding was stereoselective since neither leukotriene B4 (LTB4), lipoxin B4 (LXB4), (6S)-LXA4, 11-trans-LXA4, nor SKF 104353 competed for [3H]LXA4-specific binding while LTD4 and LTC4 partially competed. Subcellular fractionation revealed that specific binding with [3H]LXA4 was associated with membrane (42.1%)-, granule (34.5%)-, and nuclear (23.3%)-enriched fractions, a distribution distinct from that of [14,15-3H] LTB4 binding. [11,12-3H]LXA4-specific binding was modulated by guanosine analogs, suggesting the involvement of G proteins. A fluorescent LXA4 derivative (methyl-7-methoxycoumarin-LXA4) competed with [3H]LXA4 binding to intact PMN and showed specific and reversible binding as monitored by flow cytometric analysis. These results indicate that PMN possess specific recognition sites for LXA4 which may mediate its actions.

Binding Sites↗
PubMed1992

Blocking the interleukin-1 receptor inhibits leukotriene B4 and prostaglandin E2 generation in human monocyte cultures.

Interleukin-1 is a potent stimulator of arachidonic acid (AA) metabolism and this activity could be attributed to the activation of the prostaglandin-forming enzyme cyclooxygenase or of the arachidonic-releasing enzyme phospholipase A2 or both. Prostaglandin E2 (PGE2), a cyclooxygenase product, and LTB4 (5-(S),12-(R)-dihydroxy-6,14-cis-8,10-trans-eicosatetraenoic acid), a lipoxygenase product, are potent mediators of inflammation. Recently a new cytokine produced by macrophages and named interleukin-1 receptor antagonist (IL-1ra) (MW 22,000 Da) which specifically binds and blocks IL-1 receptors, has proven to be a potent inflammatory inhibitor. In our studies we found that monocyte suspensions, pretreated with hrIL-1ra at increasing concentrations (0.25-250 ng/ml) for 10 min and then treated with LPS in an overnight incubation inhibits, in a dose-dependent manner, the generation of LTB4 as measured by the highly sensitive radioimmunoassay method. In monocytes pretreated with hrIL-1ra (250 ng/ml) for 10 min and treated with arachidonic acid (10(-5)-10(-9) M) and LPS overnight, the release of LTB4 was partially inhibited when compared to hrIL-1ra-untreated cells. Moreover, hrIL-1ra (250 ng/ml) caused a partial inhibition of monocyte LTB4 production when the cells were activated with AA (10(-7) M) and then treated with IL-1 beta (5 ng/ml) overnight or 24 hr incubation. In addition, human monocytes pretreated for 10 min with increasing doses of hrIL-1ra (0.25-250 ng/ml) and then treated with hrIL-1 alpha (5 ng/ml) or beta (5 ng/ml) for 18 hr, also resulted in the inhibition of PGE2 generation as measured by RIA when compared with hrIL-1ra-untreated cells. When the cells were treated with hrIL-1ra (250 ng/ml) and activated for 18 and 48 hr with increasing doses of hrIL-1 beta a strong inhibitory effect was found on PGE2 production. HrIL-1ra used at 15 ng/ml gave a partial inhibition of LTB4 generation, after LPS (1-100 ng/ml) treatment, while NDGA totally blocked the production of LTB4. Moreover, PGE2 released by macrophages activated with LPS (100 ng/ml) or hrIL-1 beta (5 ng/ml) at 18 hr incubation time was strongly inhibited when hrIL-1ra (250 ng/ml) was used. These data suggest that the inhibition of LTB4 and PGE2 by this new macrophage-derived monokine IL-1ra occurs through the block of the IL-1 receptor, rather than phospholipase A2, and thus IL-1ra may offer a potential therapeutic approach to inflammatory states.

Arachidonic Acid↗
PubMed1992

Granulocyte-macrophage colony stimulating factor potentiates human polymorphonuclear leukocyte aggregation responses to formyl-methionyl-leucyl-phenylalanine.

Polymorphonuclear leukocytes (PMN) are known to be activated by several lymphokines and can be induced to release lysosomal enzymes, prostaglandins (PG), thromboxanes (TX) and lipoxygenase products that may be involved in PMN aggregation responses during inflammatory reactions. Granulocyte-macrophage colony stimulating factor (GM-CSF), a glycoprotein cytokine released by immunocompetent cells, has been found to prime neutrophil responses, such as increased cell aggregation after exposure to various biological stimulants. In this study, we examined the effects of the cytokine GM-CSF on human neutrophilic aggregation stimulated by N-formyl-methionyl-leucyl-phenylalanine (FMLP) and its influence on the production of various arachidonic acid metabolites. Neutrophil aggregation of purified PMNs was measured by the percent change in light transmission in a standard aggregometer, and the arachidonic acid products leukotriene B4 (LTB4) and thromboxane A2 (TXA2) were quantified by radioimmunoassay. We found that GM-CSF and other cytokines, used alone, did not cause any significant increase in aggregation of the PMN. However, prior exposure of PMN to GM-CSF markedly increased the aggregation induced by FMLP as opposed to that detected with PMN stimulated with only FMLP. This priming effect was not observed with PMN preincubated with interleukin-1 (IL-1), tumor necrosis factor (TNF) or interleukin-6 (IL-6). In addition, GM-CSF and IL-6 both failed to stimulate the production of LTB4 and TXA2, products which are known to induce PMN aggregation. These findings provide new evidence suggesting that GM-CSF facilitates the action of FMLP on the adhesion dependent cellular functions of the inflammatory response, serving as an important co-factor in neutrophil aggregation.

Calcimycin↗
PubMed1990

Formation of lipoxins and leukotrienes during receptor-mediated interactions of human platelets and recombinant human granulocyte/macrophage colony-stimulating factor-primed neutrophils.

The generation of lipoxygenase products of arachidonic acid is considered an important event in inflammation. This study demonstrates the levels of both lipoxins and leukotrienes (LTC4, LTD4, LTB4, and omega-oxidized LTB4) generated from endogenous sources of arachidonate by PMN primed with recombinant human granulocyte/macrophage colony-stimulating factor and in coincubations with platelets (1:1 to 1:100 ratio). Upon exposure to receptor-mediated stimuli (FMLP and thrombin), the levels of lipoxins generated were within the range of both LTB4 and LTC4. Co-incubation of [1-14C]arachidonate-labeled platelets with primed polymorphonuclear leukocytes (PMN) followed by addition of thrombin and FMLP led to the formation of both 5- and 15-LO products that carried 14C label. Thus, in addition to the transcellular conversion of LTA4 to platelet-derived lipoxins and LTC4, PMN can use platelet-derived arachidonate to generate lipoxygenase products. These results are the first to document the relationship between the levels of lipoxins and leukotrienes generated by receptor-mediated activation of cytokineprimed PMN interacting with platelets. Moreover, they indicate that PMN-platelet interactions utilize bidirectional transcellular routes to contribute to lipoxin formation.

Arachidonic Acids↗
PubMed1990

Evaluation of synthetic sphingosine, lysosphingolipids and glycosphingolipids as inhibitors of functional responses of human neutrophils.

Human neutrophils, when exposed to soluble stimuli, aggregate, release oxygenated products of arachidonic acid and generate active oxygen species. Sphingolipid-derived products such as sphingosine and lysosphingolipids have been shown to exert selective actions on a variety of cell types, including neutrophils. Therefore, to determine the structural basis for selective inhibition of neutrophil responses by naturally occurring sphingolipids, seven compounds were prepared by total organic synthesis, and their impact on neutrophils in suspension has been studied. The compounds synthesized included sphingosine, psychosine, lactosyl lysosphingolipid, globotriaosyl (Gb3) lysosphingolipid, galactosyl cerebroside, lactosyl ceramide and Gb3 ceramide. The neutrophil responses studied were aggregation, leukotriene generation and superoxide anion production. When exposed to non-cytotoxic levels of the synthetic compounds, as monitored by exclusion of Trypan Blue, none of the synthetic sphingolipids inhibited A23187-induced aggregation of neutrophils. Only lactosyl lysosphingolipid, at a concentration of 1 microM, significantly inhibited aggregation induced by fMetLeuPhe; the other compounds in this series including sphingosine were without effect at equal molar concentrations (1 microM). Aggregation induced by phorbol 12-myristate 13-acetate (PMA) (0.1 microM) was significantly blocked by only two of the synthetic sphingolipids (1 microM). At concentrations below 1 microM, these inhibitory actions were not evident, nor was it possible to assign a structure-activity relationship for this series of compounds. None of the synthetic sphingolipids effectively inhibited the generation of superoxide anions induced by PMA. In addition, neither synthetic sphingosine nor psychosine affected either the formation or metabolism of leukotriene B4. Taken together, the results provide further evidence that sphingolipids, when added to intact cells, are not potent selective inhibitors of functional responses of human neutrophils.

Antigens, CD↗
PubMed1990

Lipoxin A4 and lipoxin B4 stimulate the release but not the oxygenation of arachidonic acid in human neutrophils: dissociation between lipid remodeling and adhesion.

The profiles of actions of lipoxin A4 (LXA4) and lipoxin B4 (LXB4), two lipoxygenase-derived eicosanoids, were examined with human neutrophils. At nanomolar concentrations, LXA4 and LXB4 each stimulated the release of [1-14C]arachidonic acid from esterified sources in neutrophils. Lipoxin-induced release of [1-14C]arachidonic acid was both dose- and time-dependent and was comparable to that induced by the chemotactic peptide f-met-leu-phe. Time-course studies revealed that lipoxin A4 and lipoxin B4 each induced a biphasic release of [1-14C]arachidonic acid, which was evident within seconds (5-15 sec) in its initial phase and minutes (greater than 30 sec) in the second phase. In contrast, the all-trans isomers of LXA4 and LXB4 did not provoke [1-14C]AA release. Lipoxin-induced release of arachidonic acid was inhibited by prior treatment of the cells with pertussis toxin but not by its beta-oligomers, suggesting the involvement of guaninine nucleotide-binding regulatory proteins in this event. Dual radiolabeling of neutrophil phospholipid classes with [1-14C]arachidonic acid and [3H]palmitic acid showed that phosphatidylcholine was a major source of lipoxin-induced release of [1-14C]arachidonic acid. They also demonstrated that lipoxins rapidly stimulate both formation of phosphatidic acid as well as phospholipid remodeling. Although both LXA4 and LXB4 (10(-8)-10(-6) M) stimulated the release of [1-14C]arachidonic acid, neither compound evoked its oxygenation by either the 5- or 15-lipoxygenase pathways (including the formation of LTB4, 20-COOH-LTB4, 5-HETE, or 15-HETE). LXA4 and LXB4 (10(-7) M) each stimulated the elevation of cytosolic Ca2+ as monitored with Fura 2-loaded cells, albeit to a lesser extent than equimolar concentrations of FMLP. Neither lipoxin altered the binding of [3H]LTB4 to its receptor on neutrophils. In addition, they did not stimulate aggregation or induce adhesion of neutrophils to human endothelial cells. Results indicate that both LXA4 and LXB4 stimulate the rapid remodeling of neutrophil phospholipids to release arachidonic acid without provoking either aggregation or the formation of lipoxygenase-derived products within a similar temporal and dose range. Together they indicate that LXA4 and LXB4 display selective actions with human neutrophils and suggest that these eicosanoids possess unique profiles of action which may regulate neutrophil function during inflammation.

Arachidonic Acids↗
PubMed1990

Leukocyte inhibitory factor activates human neutrophils and macrophages to release leukotriene B4 and thromboxanes.

Recent evidence has proved that cytokines can stimulate the production of 5-lipoxygenase products. Leukotriene B4 (LTB4) is a major mediator of leukocyte activation in acute inflammatory reactions, which produce chemotaxis, lysosomal enzyme release, and cell aggregation. Leukocyte inhibitory factor (LIF) also causes biological responses related to inflammation, i.e., LIF directly induces specific granule secretion by polymorphonuclears (PMNs) and potentiates many formyl-methionyl-leucyl-phenylalanine (FMLPs) mediated responses. Since arachidonic acid products are important mediators of inflammation, we have studied the effects of LIF on the arachidonic acid cascade products LTB4 and thromboxane A2 (TxA2). Resuspended at a final concentration of greater than 95% polymorphonuclear PMNs were isolated and tested with some cytokines on the release of LTB4 and TxA2. Peripheral blood mononuclear cells were isolated and seeded in Petri dishes and incubated for 60 min. Adherent macrophages were used for the cytokine stimulation study. Both types of leukocytes were treated with LIF, interleukin 6 (IL 6), and granulocyte-monocyte colony stimulating factor (GM-CSF) at different concentrations, and test agents A23187 and FMLP. Radioimmunoassay for LTB4 and TxB2 was determined by the resulting supernatants. Treatment of PMNs and macrophages with LIF at different concentrations proved to generate significant increases in LTB4 and TxA2 production. This was compared with IL 6 and GM-CSF, which had no effects. In these experiments, TxA2 generations could not be attributed to platelet contamination of PMN suspensions. The quantity of platelet contamination was not sufficient to influence how much TxB2 was produced. The similarities of LIF to other arachidonate stimulating cytokines suggest a similar mode of action in producing hematologic changes typical of tissue injury.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcimycin↗
PubMed1989

Phospholipid bilayers enhance the stability of leukotriene A4 and epoxytetraenes: stabilization of eicosanoids by liposomes.

The hydrolysis of leukotriene A4 and two epoxytetraenes was examined in the presence and absence of liposomes. When added to liposomes in suspension, the stability of LTA4 was increased in a time- and dose-dependent fashion. At 10 min, the half-life of LTA4 was increased 67.1 +/- 6.8% in the presence of liposomes which was comparable to that observed with albumin (10 mg/ml): 68.3 +/- 6.9%. Phosphatidylcholine, in a non-bilayer configuration, was also effective in enhancing the half-life of LTA4, albeit to a lesser extent than liposomes. At equal molar concentrations, the enhanced stability of eicosanoid epoxides with liposomes gave a rank order with leukotriene A4 greater than 5(6)epoxytetraene greater than 14(15)epoxytetraene. Results indicate that phospholipid bilayers can protect leukotriene A4 and 5(6)epoxytetraene from non-enzymatic hydrolysis. Moreover, they suggest that the biological half-life of intermediates involved in the formation of both leukotrienes and lipoxins can be increased by their association with membranes.

Albumins↗