Aberrant production of leukotriene C4 by macrophages from autoimmune-prone mice.

Eicosanoids have been implicated in the pathogenesis of autoimmune diseases. In this study, peritoneal macrophages from autoimmune-prone mice were examined for their capacity to produce proinflammatory 5-lipoxygenase metabolites. The results indicate that enhanced production of leukotriene C4 is a common feature of murine autoimmunity and suggest further that aberrations in 5-lipoxygenase activity may play a role in the development of lupus.

Stimulus specificity of the generation of leukotrienes by dog mastocytoma cells.

Isolated dog mastocytoma cells sensitized with dog anti-ragweed IgE and challenged with ragweed antigen or incubated with ionophore A23187 or the carboxy-terminal dodecapeptide of platelet factor 4, PF4(59-70), release histamine and concurrently generate leukotrienes B4, C4, and D4. In contrast, the exposure of mastocytoma cells to 0.1-3 micrograms/ml of 15-hydroxyeicosatetraenoic acid (15-HETE) stimulates selectively the generation of leukotrienes, in the absence of histamine release, while 0.1-1 micrograms/ml of compound 48/80 releases histamine without enhancing the generation of leukotrienes. That natural stimuli are capable of selectively activating one synthetic or secretory compartment of mast cells suggests that separate subsets of receptors as well as different biochemical events may serve to mobilize each class of mediators.

Formation of slow-reacting substance of anaphylaxis in human lung tissue and cells before release.

The capacity to extract slow-reacting substance of anaphylaxis (SRS-A) from human lung tissue or cells after immunologic activation, together with the measurement of SRS-A in both the extract and the surrounding fluid, permits study of total SRS-A generation. That the material extracted is SRS-A was established by both differential bioassay and purification. SRS-A accumulation was entirely intracellular after limited IgE-dependent direct or reversed anaphylactic activation. Intracellular accumulation also generally preceded release, with generation of SRS-A continuing well beyond a plateau in the cellular SRS-A level and the release of preformed mediators. The quantity of SRS-A generated after immunologic activation was modulated by the introduction of exogenous cyclic nucleotides, revealing a site of cyclic nucleotide action distinct from that on mediator release. The capacity to determine not only the release of preformed mediators but also the generation of a newly formed mediator, the sum of SRS-A in cells and supernate, adds an additional dimension to the analysis of the cellular events of immediate hypersensitivity.

Leukotriene generation by eosinophils.

Horse eosinophils purified to greater than 98% generated slow reacting substance (SRS) when incubated with the calcium ionophore A23187. On a per cell basis, eosinophils generated four to five times the SRS produced by similarly treated horse neutrophils. Eosinophil SRS production was inhibited by 5,8,11,14-eicosatetraynoic acid and augmented by indomethacin and arachidonic acid, suggesting that it was a product(s) of the lipoxygenase pathway of arachidonic acid metabolism. Compounds with SRS activity were purified by high-pressure liquid chromatography (HPLC) and identified by ultraviolet spectra, spectral shift on treatment with lipoxygenase, incorporation of [14C]arachidonic acid, gas chromatography-mass spectrometry, and comparison of retention times on HPLC to authentic standards. The eosinophil products characterized were 5-(S), 12-(R)-dihydroxy-6-cis-8, 10-trans-14-cis-eicosatetraenoic acid (leukotriene B4) and its 5-(S), 12-(R)-6-trans and 5-(S), 12-(S)-6-trans isomers, 5-(S)-hydroxy-6-(R)-S-glutathionyl-7,9-trans-11, 14-cis-eicosatetraenoic acid (leukotriene C4) and its 11-trans isomer, and 5-(S)-hydroxy-6-(R)-S-cysteinylglycine-7,9-trans-11,14-cis-eicosatetraenoic acid (leukotriene D4).

Cyclooxygenase blockade elevates leukotriene E4 production during acute anaphylaxis in sheep.

We examined changes in the levels of eicosanoids in blood and pulmonary lymph of anesthetized sheep undergoing acute anaphylaxis. Within 1-3 min of intravenous antigenic challenge of previously sensitized sheep, there were approximately 7-30-fold elevations in mean arterial plasma levels of thromboxane B2 and 6-ketoprostaglandin F1 alpha, respectively, as measured by RIA. Negligible changes in levels of these cyclooxygenase products were found in both nonsensitized sheep and in sensitized sheep treated with indomethacin before antigenic challenge. In contrast, no changes in levels of sulfidopeptide leukotrienes (SPLT) in pulmonary lymph were detectable by RIA during anaphylaxis in sensitized or nonsensitized sheep, but levels of SPLT in indomethacin-treated sensitized sheep increased more than fivefold above levels in lymph from both other groups of animals. The immunoreactive SPLT in lymph from indomethacin-treated sheep was accounted for as LTE4, as demonstrated by mobility on HPLC and absorbance at 280 nm. These results support the possibility that certain undesirable effects of nonsteroidal antiinflammatory drugs, such as cardiopulmonary reactions in aspirin-sensitive individuals, and impaired renal and cardiac function during therapy with these drugs, may be related in part to augmented synthesis of the 5-lipoxygenase pathway products, especially those of the sulfidopeptide class. Increased LT production could also limit the antiinflammatory effectiveness of these drugs in many disease states.

Synthesis of leukotriene C and other arachidonic acid metabolites by mouse pulmonary macrophages.

Mouse resident pulmonary macrophages were subdivided into alveolar (PAM) and interstitial (PTM) populations on the basis of accessibility to pulmonary lavage, and zymosan-induced arachidonic acid (20:4) metabolism was examined in both populations labeled with [3H]20:4. Maximal phagocytic doses of unopsonized zymosan induced the specific release of 11% of phospholipid 20:4 by PTM and 4.6% by PAM. Direct fatty acid analysis of [3H]20:4-labeled PTM cultured in the presence or absence of zymosan indicated that the specific activity of the [3H]20:4 in cell phospholipid provided an accurate measure of 20:4 released by the cells, and could therefore be used to quantitate the synthesis of 20:4 metabolites by PTM in vitro. The single major 20:4 metabolite of PTM was the slow-reacting substance leukotriene C, which was synthesized in quantities of 3-4 pmol/microgram cell protein (280-370 pmol/10(6) cells), and comprised 20-25% of the released 20:4. PTM also synthesized prostaglandin E2, prostacyclin, thromboxane A2, and hydroxyeicosatetraenoic acids. In contrast, PAM produced leukotrienes D and E in addition to leukotriene C, prostaglandin E2, thromboxane A2, and hydroxyeicosatetraenoic acids. Prostacyclin formation by PAM was not observed. These studies define a set of experimental conditions for the study of 20:4 metabolism by pulmonary macrophages, and demonstrate that these cells are rich sources of LTC as well as other 20:4 oxygenated products.

Regulation of human eosinophil viability, density, and function by granulocyte/macrophage colony-stimulating factor in the presence of 3T3 fibroblasts.

Normodense human peripheral blood eosinophils were isolated under sterile conditions from the 22/23 and 23/24% interfaces and the cell pellet of metrizamide gradients. After culture for 7 d in RPMI media in the presence of 50 pM biosynthetic (recombinant) human granulocyte/macrophage colony-stimulating factor (rH GM-CSF), 43 +/- 7% (mean +/- SEM, n = 8) of the cells were viable; in the absence of rH GM-CSF, no eosinophils survived. The rH GM-CSF-mediated viability was concentration dependent; increased survival began at a concentration of 1 pM, a 50% maximal response was attained at approximately 3 pM, and a maximal effect was reached at concentrations of greater than or equal to 10 pM rH GM-CSF. In the presence of rH GM-CSF and mouse 3T3 fibroblasts, 67 +/- 6% (mean +/- SEM, n = 8) of the eosinophils survived for 7 d. In a comparative analysis, there was no difference in eosinophil viability after 7 and 14 d (n = 3) in the presence of 50 pM GM-CSF and fibroblasts. Culture with fibroblasts alone did not support eosinophil survival. The addition of fibroblast-conditioned media to rH GM-CSF did not further improve eosinophil viability, indicating a primary role for GM-CSF in supporting these eosinophil cell suspensions ex vivo and a supplementary role for 3T3 fibroblasts. Eosinophils cultured for 7 d localized on density gradient sedimentation at the medium/18, 18/20, and 20/21 interfaces of metrizamide gradients, indicating a change to the hypodense phenotype from their original normodense condition. In addition, the cultured eosinophils generated approximately 2.5-fold more LTC4 than freshly isolated cells when stimulated with the calcium ionophore A23187 and manifested sevenfold greater antibody-dependent killing of S. mansoni larvae than the freshly isolated, normodense cells from the same donor. Thus we demonstrate the rH GM-CSF dependent conversion in vitro of normodense human eosinophils to hypodense cells possessing the augmented biochemical and biological properties characteristic of the hypodense eosinophils associated with a variety of hypereosinophilic syndromes. In addition, these studies provide a culture model of at least 14 d suitable for the further characterization of hypodense eosinophils.

Prevention of endogenous leukotriene production during anaphylaxis in the guinea pig by an inhibitor of leukotriene biosynthesis (MK-886) but not by dexamethasone.

Leukotriene C4 (LTC4) underwent rapid elimination from the circulating blood and was extensively converted to LTD4 within the vascular space of the guinea pig. To mimic the elimination and metabolism of endogenous LTC4 generated during anaphylaxis, 14,15-3H-labeled LTC4 was infused intravenously over a period of 15 min, leading to a recovery in bile of 85% of the infused LT radioactivity within 2 h. Corresponding to the tracer studies, LTD4 and, to a lesser extent, LTC4 were the predominant endogenous cysteinyl LTs in guinea pig bile. The biliary production rate of endogenous LTD4 increased from 0.3 +/- 0.1 to 6.2 +/- 1.8 pmol x min-1 x kg-1 (p less than 0.001) during anaphylactic shock induced by intravenous injection of OVA (0.2 mg/kg) into sensitized guinea pigs. A novel LT biosynthesis inhibitor (MK-886; 10 mg/kg, i.v., 15 min before antigen challenge) suppressed the antigen-induced cysteinyl LT production by greater than 92% (p less than 0.001). This inhibition of systemic LTC4 formation was associated with a complete protection against lethal anaphylactic shock in animals pretreated in addition with the H1 receptor antagonist pyrilamine. Pretreatment with either the inhibitor of LT synthesis or the histamine receptor antagonist reduced the lethality during anaphylactic shock from 100 to 60 and 78%, respectively. In artificially ventilated, pyrilamine-pretreated animals, the antigen-induced decrease in dynamic lung compliance and the rise in hematocrit were significantly reduced (p less than 0.05) by pretreatment with the inhibitor of LT synthesis. Dexamethasone at high doses (10 mg/kg, i.p., once daily for 7 d, or in a single dose of 10 mg/kg, i.v., 3.5 h before challenge) had no inhibitory effect on LT generation during anaphylaxis in vivo. However, in resident peritoneal macrophages, harvested from these dexamethasone-treated sensitized guinea pigs and stimulated with zymosan, both cysteinyl LT and 6-keto-PGF1 alpha formation were strongly suppressed. These studies indicate an important role of cysteinyl LTs in systemic anaphylaxis in vivo and demonstrate the blockade of anaphylactic LT generation by a novel inhibitor of LT biosynthesis (MK-886) but not by dexamethasone.

RANTES and macrophage inflammatory protein 1 alpha induce the migration and activation of normal human eosinophil granulocytes.

The cellular infiltrates of certain inflammatory processes found in parasitic infection or in allergic diseases consist predominantly of eosinophilic granulocytes, often in association with activated T cells. This suggests the existence of chemotactic agonists specific for eosinophils and lymphocyte subsets devoid of neutrophil-activating properties. We therefore examined four members of the intercrine/chemokine superfamily of cytokines (monocyte chemotactic peptide 1 [MCP-1], RANTES, macrophage inflammatory protein 1 alpha [MIP-1 alpha], and MIP-1 beta), which do not activate neutrophils, for their ability to affect different eosinophil effector functions. RANTES strongly attracted normal human eosinophils by a chemotactic rather than a chemokinetic mechanism with a similar efficacy as the most potent chemotactic myeloid cell agonist, C5a. MIP-1 alpha also induced eosinophil migration, however, with lower efficacy. RANTES and MIP-1 alpha induced eosinophil cationic protein release in cytochalasin B-treated eosinophils, but did not promote leukotriene C4 formation by eosinophils, even after preincubation with interleukin 3 (IL-3), in contrast to other chemotactic agonists such as C5a and formyl-methionyl-leucyl-phenylalanine (FMLP). RANTES, but not MIP-1 alpha, induced a biphasic chemiluminescence response, however, of lower magnitude than C5a. RANTES and MIP-1 alpha both promoted identical transient changes in intracellular free calcium concentration ([Ca2+]i), with kinetics similar to those induced by chemotactic peptides known to interact with G protein-coupled receptors. No cross-desensitization towards other peptide agonists (e.g., C5a, IL-8, FMLP) was observed, suggesting the presence of specific receptors. Despite its weaker eosinophil-activating properties, MIP-1 alpha was at least 10 times more potent on a molar basis than RANTES at inducing [Ca2+]i changes. Interestingly, RANTES deactivated the MIP-1 alpha-induced [Ca2+]i changes, while the RANTES response was preserved after MIP-1 alpha stimulation. MCP-1, a potent monocyte chemoattractant and basophil agonist, as well as MIP-1 beta, a peptide with pronounced homology to MIP-1 alpha, did not activate the eosinophil functions tested. Our results indicate that RANTES and MIP-1 alpha are crucial mediators of inflammatory processes in which eosinophils predominate.

Synthesis of compounds with properties of leukotrienes C4 and D4 in gerbil brains after ischemia and reperfusion.

6-Sulfidopeptide-containing leukotriene-like immunoreactivity was synthesized in gerbil forebrains after bilateral common carotid occlusion and reperfusion. At 5, 10, or 15 minutes of ischemia, concentrations increased significantly and became more marked on reperfusion. Immunoreactivity was highest in forebrain gray matter and was below the detection limit of the assay in brain regions remote from the zone of ischemia. In vitro experiments with vascular cells and organ cultures of cerebral arteries indicate that the cerebral blood vessel wall is not a major source of biosynthetic activity in the brain. These experiments demonstrate leukotriene biosynthesis by the brain. Because synthesis occurs during ischemia and reperfusion and because leukotrienes are potent vasoconstrictors and promoters of tissue edema, they may play a role in the pathophysiology of cerebral ischemia.

Leukotrienes and lipoxins: structures, biosynthesis, and biological effects.

Arachidonic acid is released from membrane phospholipids upon cell stimulation (for example, by immune complexes and calcium ionophores) and converted to leukotrienes by a 5-lipoxygenase that also has leukotriene A4 synthetase activity. Leukotriene A4, an unstable epoxide, is hydrolyzed to leukotriene B4 or conjugated with glutathione to yield leukotriene C4 and its metabolites, leukotriene D4 and leukotriene E4. The leukotrienes participate in host defense reactions and pathophysiological conditions such as immediate hypersensitivity and inflammation. Recent studies also suggest a neuroendocrine role for leukotriene C4 in luteinizing hormone secretion. Lipoxins are formed by the action of 5- and 15-lipoxygenases on arachidonic acid. Lipoxin A causes contraction of guinea pig lung strips and dilation of the microvasculature. Both lipoxin A and B inhibit natural killer cell cytotoxicity. Thus, the multiple interaction of lipoxygenases generates compounds that can regulate specific cellular responses of importance in inflammation and immunity.

Nonimmunological production of leukotrienes induced by platelet-activating factor.

Platelet-activating factor caused rapid pulmonary vasoconstriction and edema in isolated lungs perfused with albumin-free salt solution devoid of formed blood elements. These effects may be due in part to the action of leukotrienes D4 and C4, which were identified by bioassay and high-pressure liquid chromatography in the lung effluent after stimulation by platelet-activating factor. These findings help illuminate some of the deleterious effects that platelet-activating factor elicits in anaphylactic reactions and possibly in other forms of lung injury.

Eosinophils cocultured with endothelial cells have increased survival and functional properties.

Human peripheral blood eosinophils, cells often associated with allergic and parasitic diseases, were maintained in vitro for at least 14 days when they were cocultured with bovine endothelial cells and for at least 7 days when cultured with either bovine or human endothelial cell-derived conditioned medium. The cocultured eosinophils became hypodense and generated about three times as much leukotriene C4 upon activation with calcium ionophore and killed about three times as many antibody-coated larvae of Schistosoma mansoni as freshly isolated normodense eosinophils. That these cells can be maintained in vitro by coculture with endothelial cells, and the surprising finding that the cocultured eosinophils have biochemical, cytotoxic, and density properties similar to those of eosinophils in patients with allergic and other disorders, will facilitate investigation of the regulation and role of these cells in health and disease.

Leukotrienes as mediators in tissue trauma.

A significant increase in the production of cysteinyl leukotrienes was observed after mechanical or thermal trauma in the anesthesized rat. The amount of biliary N-acetyl-leukotriene E4, which represents a suitable indicator for blood plasma leukotrienes, was used as a measure of leukotriene generation. Cysteinyl leukotrienes were rapidly eliminated from blood plasma into bile where N-acetyl-leukotriene E4 was the major metabolite. Leukotrienes were at a much lower concentration in blood plasma than in bile and differed in the pattern of metabolites. The detected amounts of leukotrienes were sufficient to induce known phenomena associated with trauma, such as tissue edema and circulatory and respiratory dysfunction. Increased leukotriene generation appears to play an important role in the pathophysiology of tissue trauma.

Eosinophils altered phenotypically and primed by culture with granulocyte/macrophage colony-stimulating factor and 3T3 fibroblasts generate leukotriene C4 in response to FMLP.

Normodense eosinophils failed to generate leukotriene C4 (LTC4) in response to incremental concentrations of FMLP but did produce LTC4 when stimulated with calcium ionophore A23187. Normodense eosinophils, maintained in culture with 10(-11) M granulocyte/macrophage colony-stimulating factor (GM-CSF) in the presence of 3T3 fibroblasts, became responsive to transmembrane stimulation with FMLP by day 4 with a maximal effect by day 7. After 7 d of culture, hypodense eosinophils stimulated with 2 x 10(-7) M FMLP generated 26 ng LTC4/10(6) cells, and LTC4 biosynthesis was blocked by N-tertbutoxy-carbonyl-L-methionyl-L-leucyl-L-phenylalanine (N-t-BOC-MLP). Neither calcium ionophore stimulation of LTC4 from endogenous arachidonic acid nor substrate-initiated production of LTC4 from incorporated LTA4 changed when eosinophils were cocultured with GM-CSF and 3T3 fibroblasts. Furthermore, when incubated with 10(-6) M FMLP, normodense eosinophils generated no net superoxide measured by the reduction of cytochrome c, whereas replicate eosinophils cultured for 7 d with 10(-11) M GM-CSF and 3T3 fibroblasts reduced a net of 17 nmol of cytochrome c/10(6) cells. These studies suggest that primed and phenotypically altered eosinophils present at an extravascular site may exert pathobiologic effects by responding to soluble ligands in the tissues.

Thromboxane synthase is preferentially conserved in activated mouse peritoneal macrophages.

Resident macrophages isolated from uninfected animals produce large quantities of arachidonic acid (AA) metabolites. Immunizing animals with protein antigens or bacteria activates macrophages and causes an 80% reduction in the cyclooxygenase and lipoxygenase metabolites relative to resident cells. Since some products have been shown to modulate immune functions, we examined how the AA metabolic enzyme activities regulate the products that are synthesized. We demonstrate that the cyclooxygenase, 5-lipoxygenase, prostacyclin synthase, and probably prostaglandin (PG) endoperoxide E-isomerase activities were decreased in activated peritoneal macrophages. In sharp contrast, thromboxane synthase activity was selectively unchanged or enhanced in the activated macrophages. Thus the immune response appears to modulate the activity of the AA and PG endoperoxide-dependent enzymes, thus dictating a major shift in the profile of metabolites synthesized by macrophages.

Synthesis of hydroxyeicosatetraenoic acids and leukotrienes in rat nephrotoxic serum glomerulonephritis. Role of anti-glomerular basement membrane antibody dose, complement, and neutrophiles.

The basal and stimulated synthesis of immunoassayable 12- and 5-monohydroxyeicosatetraenoic acids (HETE) and leukotrienes (LT) B4 and C4 was studied in glomeruli isolated from rats with nephrotoxic serum glomerulonephritis (NSGN) induced by low (30 micrograms/g body weight) or high (105 micrograms/g) doses of anti-rat glomerular basement membrane (GBM) immunoglobulin (Ig). In the early heterologous phase of the disease, low doses of anti-GBM Ig enhanced the basal synthesis of 12-HETE but not that of 5-HETE or LT. High anti-GBM Ig doses enhanced the basal synthesis of 5-HETE and LTB4 as well. Under stimulated conditions, enhanced glomerular production of 5-HETE and LTB4 occurred at 15 min after infusion of anti-GBM Ig, peaked at 1 h, and returned toward control levels by 24 h. At 48 h, 72 h, and on day 12, the synthesis of these eicosanoids was impaired. Neutrophile depletion only partially reduced glomerular eicosanoid synthesis after induction of NSGN whereas complement depletion significantly reduced 5-HETE, 12-HETE, and LTB4. These observations indicate that in the heterologous phase of NSGN there is enhanced but short-lived glomerular 5-HETE and LTB4 synthesis. This phenomenon is mediated by complement activation and may be an important proinflammatory event leading to capillary wall injury in the early stages of the disease.

Human eosinophils have prolonged survival, enhanced functional properties, and become hypodense when exposed to human interleukin 3.

Human eosinophils were cultured in the presence of recombinant human IL-3 for up to 14 d and their biochemical, functional, and density properties were assessed. After 3 d of culture in 10 pM IL-3, eosinophils had a viability of 70% compared with only 10% in enriched medium alone. Neither IL-1 alpha, IL-2, IL-4, tumor necrosis factor, basic fibroblast growth factor, nor platelet-derived growth factor maintained eosinophil viability. The 7- and 14-d survival of the cultured eosinophils was 55 and 53%, respectively. No other cell type, including neutrophils, was present after culture. After 7 d of culture, the normodense eosinophils were converted to hypodense cells as assessed by density centrifugation. Eosinophils exposed to 1,000 pM IL-3 for 30 min or cultured in 10 pM IL-3 for 7 d generated approximately threefold more leukotriene C4 (LTC4) in response to calcium ionophore than freshly isolated cells. Furthermore, whereas freshly isolated eosinophils killed only 14% of the antibody-coated Schistosoma mansoni larvae, these eosinophils killed 54% of the larvae when exposed to 100 pM IL-3. The enhanced helminth cytotoxicity was maintained for 7 d when eosinophils were cultured in the presence of both 10 pM IL-3 and 3T3 fibroblasts, but not when eosinophils were cultured in the presence of IL-3 alone. IL-3 thus maintains the viability of eosinophils in vitro, augments the calcium ionophore-induced generation of LTC4, enhances cytotoxicity against antibody-sensitized helminths, and induces the eosinophils to become hypodense cells. These phenotypic changes in the eosinophil may be advantageous to host defense against helminthic infections but may be disadvantageous in conditions such as allergic disease.

Stimulated arachidonate metabolism during foam cell transformation of mouse peritoneal macrophages with oxidized low density lipoprotein.

Changes in arachidonate metabolism were examined in mouse peritoneal macrophages incubated with various types of lipoproteins. Oxidized low density lipoprotein (LDL) was incorporated by macrophages and stimulated macrophage prostaglandin E2 (PGE2) and leukotriene C4 syntheses, respectively, 10.8- and 10.7-fold higher than by the control. Production of 6-keto-PGF1 alpha, a stable metabolite of prostacyclin, was also stimulated. No stimulation was found with native LDL, which was minimally incorporated by the cells. Acetylated LDL and beta-migrating very low density lipoprotein (beta-VLDL), though incorporated more efficiently than oxidized LDL, also had no stimulatory effect. When oxidized LDL was separated into the lipoprotein-lipid peroxide complex and free lipid peroxides, most of the stimulatory activity was found in the former fraction, indicating that stimulation of arachidonate metabolism in the cell is associated with uptake of the lipoprotein-lipid peroxide complex. These results suggest that peroxidative modification of LDL could contribute to the progression of atheroma by stimulating arachidonate metabolism during incorporation into macrophages.

Effects of endogenously produced leukotrienes, thromboxane, and prostaglandins on coronary vascular resistance in rabbit myocardial infarction.

In an effort to evaluate the synthesis and function of eicosanoids in myocardial infarction, we have developed a technique of in vivo myocardial infarction in rabbits followed by ex vivo cardiac perfusion. Isolated Langendorff perfused infarcted hearts (removed 1 or 4 d after infarction) responded to the inflammatory cell agonist N-formylmethionyl-leucyl-phenylalanine (fMLP) with (a) the release of leukotrienes B4, C4, and D4; (b) the release of large amounts of thromboxane (235 +/- 66 ng/5 min), prostacyclin (714 +/- 285 ng/5 min), and prostaglandin E2 (PGE2) (330 +/- 108 ng/5 min); and (c) a coronary vasoconstriction (21.1 +/- 2.5% increase in coronary perfusion pressure) that was specifically inhibited by the peptidoleukotriene receptor antagonist FPL-55712. While noninfarcted hearts challenged with fMLP also released leukotrienes B4, C4, and D4, they released only small amounts of the cyclooxygenase products (thromboxane, 30 +/- 9 ng/5 min; prostacyclin, 120 +/- 54 ng/5 min; PGE2, 27 +/- 10 ng/5 min) and showed minimal vasoconstriction (5.6 +/- 2.1% increase in perfusion pressure). Similarly, hearts challenged with fMLP 30 d following infarction released only small amounts of the cyclooxygenase products (thromboxane, 42 +/- 8 ng/5 min; prostacyclin, 386 +/- 31 ng/5 min; PGE2, 79 +/- 25 ng/5 min). When bradykinin was administered, no leukotrienes were produced, but acutely infarcted hearts released 10 times more thromboxane, prostacyclin, and PGE2 than normal hearts and significantly larger amounts of these products than 30-d infarcted hearts. Histologic analysis showed no inflammatory cells in normal hearts, a prominent polymorphonuclear leukocyte infiltration in 1-d infarcted tissue, fibroblast proliferation with mononuclear cell invasion in 4-d infarcted tissue, and a fibrotic scar with scanty mononuclear cell infiltrate in 30-d infarcted tissue. Inflammatory cell invasion was temporarily associated with augmented cyclooxygenase metabolism, suggesting that infiltrating leukocytes may be responsible for production of thromboxane, prostacyclin, and PGE2 in acutely infarcted hearts. The finding that endogenously produced peptidoleukotrienes are potent coronary vasoconstrictors in infarcted rabbit hearts suggests that these products may contribute to tissue injury in myocardial infarction.