Attilio Crea

Identification and characterization of a monocyte-derived neutrophil-activating factor in corticosteroid-resistant bronchial asthma.

Peripheral blood mononuclear cells (PBMC) were isolated from seven normal subjects, eight asthmatic subjects clinically sensitive to corticosteroids (CS), and eight asthmatic subjects clinically resistant to corticosteroids (CR). PBMC were cultured at 37 degrees C for 24 h in the absence or presence of 10(-16) to 10(-4) M hydrocortisone. Calcium ionophore (A23187)-activated neutrophils (PMN) primed by supernatants of PBMC from asthmatic subjects cultured in the absence of hydrocortisone generated approximately threefold more leukotriene B4 than PMN primed by supernatants of PBMC from normal subjects (P less than 0.05). Incubation of PBMC derived from CS subjects with 10(-8) M hydrocortisone completely inhibited the production of the enhancing activity (P less than 0.01), whereas in CR subjects hydrocortisone at concentrations up to 10(-4) M did not suppress the release of enhancing activity. The enhancing activity was produced by monocytes. Enhancing activity eluted with an Mr of 3,000 D and a pI of 7.1. It eluted at 10% acetonitrile after reverse-phase HPLC. The activity was destroyed by heating to 60 degrees C for 60 min and was sensitive to pronase treatment. The purified factor also enhanced superoxide generation by PMN which had been stimulated submaximally by phorbol myristate acetate.

Inhibition of leukotriene B4-induced neutrophil migration by lipoxin A4: Structure-function relationships

Lipoxins are trihydroxytetraene metabolites which are derived from arachidonic acid through an interaction between different lipoxygenase pathways. Previous work has shown that lipoxin A4 (LXA4) inhibits the chemotactic responsiveness of neutrophils (PMN) to leukotriene B4. We have now assessed the structural determinants of the lipoxin A4 molecule which are necessary for its inhibitory activity, using structural analogs of LXA4 prepared by chemical synthesis. Our results indicate the importance of two adjacent free hydroxyl groups in either the R or the S configuration; one hydroxyl group has to be in the C-6 position, but the other hydroxyl group can be in either the C-5 or the C-7 position for the conferment of inhibitory activity.

The metabolism of leukotrienes in blood plasma studied by high-performance liquid chromatography

The metabolism of leukotrienes (B4, C4, D4, and E4) within human plasma was studied and a simple sample preparation is presented. It was demonstrated that leukotriene E4 and leukotriene B4 were stable during incubation at 37 degrees C using the in vitro system. In contrast, leukotriene C4 was metabolized by gamma-glutamyl transpeptidase activities into leukotriene D4 which was further metabolized by dipeptidase activities of plasma into leukotriene E4. The transition state inhibitor of gamma-glutamyl transpeptidase L-serine-borate decreased the metabolism of leukotriene C4 in plasma. Dilution of plasma demonstrated that the dipeptidase was more active compared to the gamma-glutamyl transpeptidase. The metabolizing activities of plasma were functionally characterized by fractionating the plasma proteins.

Lipoxin A4 inhibits phosphoinositide hydrolysis in human neutrophils

Lipoxins (LX) are trihydroxytetraene metabolites derived from arachidonic acid via an interaction between the 5- and 15-lipoxygenases. Preincubation of [3H] myo-inositol labeled PMN with 10-7M and 10-5M LXA4 for 1 minute at 37 degrees C resulted in a concentration dependent inhibition of the generation of [3H] IP3 and [3H] IP in cells subsequently stimulated by increasing doses of LTB4 or FMLP for 1 minute at 37 degrees C. Preincubation of PMN with LXA4 did not inhibit specific binding of [3H] LTB4 to PMN. These results indicate that LXA4 inhibits chemotactic factor-induced phosphoinositide hydrolysis at a post-receptor level.

Effects of Dietary Fish Oil Lipids on Allergic and Inflammatory Diseases

Fish oil is rich in the polyunsaturated N-3 fatty acids, eicosapentaenoic (EPA) and docosahexaenoic acids (DCHA). EPA competes with arachidonic acid (AA) for metabolism by the cyclooxygenase and lipoxygenase pathways. Selective metabolites derived from EPA have reduced biological activities as compared with the AA-derived counterparts. Dietary supplementation with EPA led to incorporation of EPA into membrane phospholipids, an inhibition of 5-lipoxygenase pathway activity, and a reduction of the elaboration of platelet-activating factor. Neutrophil chemotaxis and the capacity of these cells to adhere to endothelial cells are substantially attenuated. This suggests that EPA has anti-inflammatory potential. Clinical trials in rheumatoid arthritis, psoriasis, atopic dermatitis, and bronchial asthma have shown beneficial effects. Whether the benefit obtained clinically is sufficient to replace or significantly reduce any clinical condition remains to be answered.

The contractile activities of lipoxin A4 and lipoxin B4 for guinea‐pig airway tissues

1 The isometric contractile activities of lipoxin A4 (LxA4) and lipoxin B4 (LxB4) were evaluated on guinea‐pig lung tissue over the concentration range, 10−8 to 10−5 m. 2 LxA4 contracted guinea‐pig lung parenchymal strips; the concentration eliciting 50% maximum histamine response was 3 × 10−6 m. LxA4 did not contract tracheal spirals. 3 The LxA4 dose‐response curve was parallel to that of leukotriene D4 (LTD4) with LxA4 being approximately 10,000 fold less potent than LTD4. 4 The time course of the contraction elicited by LxA4 was similar to that of LTD4 and it was slow in onset and did not plateau for 20 min. 5 Pre‐incubation of parenchymal strips with leukotriene receptor antagonists at a concentration of 1 × 10−6 m to 3 × 10−5 m FPL 55712 or 3 × 10−5 m L 649923 inhibited LxA4 activity. 6 Pre‐incubation of tissues with 1 × 10−5 m L 651392, a 5‐lipoxygenase inhibitor, or 1 × 10−5 m indomethacin, a cyclo‐oxygenase inhibitor, did not affect the contractile activity of LxA4. 7 LxB4 did not constrict parenchymal strips or tracheal spirals.

Comparison of the generation of platelet-activating factor and leukotriene C4 in human eosinophils stimulated by unopsonized zymosan and by the calcium ionophore A23187: The effects of nedocromil sodium

The generation of platelet-activating factor (PAF) and leukotriene C4 (LTC4) from normodense human eosinophils (EOSs), stimulated with unopsonized zymosan or calcium ionophore A23187, has been studied. There was a zymosan time- and dose-dependent increase in both PAF and LTC4 production. A plateau of 0.11 +/- 0.04 ng of PAF per 10(6) EOSs (mean +/- SEM; n = 7) and of 1.38 +/- 0.58 ng of LTC4 per 10(6) EOSs (n = 5) was reached at 5 x 10(8) zymosan particles at 37 degrees C for 30 minutes. Under optimal conditions, 91 +/- 1% of the PAF and 66 +/- 13% of the LTC4 remained cell associated. Calcium ionophore A23187 induced a time- and dose-dependent increase in the quantities of PAF and of LTC4 generated by EOSs. A plateau of 31 +/- 13 ng of LTC4 per 10(6) EOSs (n = 5) was reached at 1 mumol/L of calcium ionophore A23187 at 37 degrees C for 15 minutes. The dose response for PAF generation reached 4.2 +/- 0.8 ng/10(6) EOSs (n = 8) at 10 mumols/L of calcium ionophore A23187 at 37 degrees C for 15 minutes and had not plateaued; 90 +/- 5% of the generated PAF was cell associated. In vitro preincubation of EOSs with 10(-8) to 10(-4) mol/L of nedocromil sodium for 15 minutes did not change the subsequent generation or cellular distribution of PAF or LTC4 in EOSs optimally stimulated with either zymosan or calcium ionophore A23187.

Lipoxins of the 5-Series Derived from Eicosapentaenoic Acid

Arachidonic acid released from membrane phospholipids during cell activation is metabolised by the cyclo-oxygenase pathway to prostaglandins and thromboxanes and by the 5-lipoxygenase pathway to leukotrienes. In the 5-lipoxygenase pathway, arachidonic acid is metabolised through two sequential intermediates, 5-hydroperoxy eicosatetraenoic acid (5-HPETE) and leukotriene A4 (LTA4), to leukotriene B4 (LTB4) and to leukotriene C4 (LTC4) and its conversion products, leukotriene D4 (LTD4) and leukotriene E4 (LTE4).

Synthesis of 11, 12, 14, 15-tetrahydro-leukotriene C,D,E via A

Abstract The synthesis of tetrahydro-7E,9Z-leukotriene A methyl ester and its reaction with glutathione, cysteinylglycine and cysteine providing the tetrahydro analogues of leukotriene C 4 , C 4 , E 4 .

Syntheses of leukotriene analogs.

The synthesis of the C20 and C22 analogs of the leukotrienes from the unsaturated aldehydes 2a, b with undecyl(triphenyl)phosphorane and reactions with thiopeptides providing the conjugates LTC, LTD, and LTE is described.