Tolfenamic acid inhibits leukotriene synthesis in human granulocytes

Conclusions Higher concentrations of TA were needed to inhibit chemiluminescence than LTB₄ formation (whereas indomethacin and timegadine reduced these two phenomena with similar potencies) suggesting a specific blockade of 5-lipoxygenase by TA. Oral TA decreased leukotriene production in PMNsex vivo. This may be related to the good anti-inflammatory and anti-migraine actions of TA, and partly explain its rare gastric and bronchoconstrictive adverse effects. However, the recommended single oral dose (100–400 mg) seems to be too low to decrease systemic LT-synthesis in all subjects.     

Activated factor XI inhibits chemotaxis of polymorphonuclear leukocytes

PMN leukocytes are the most abundant leukocytes in the circulation and play an important role in host defense. PMN leukocyte recruitment and inflammatory responses at sites of infection are critical components in innate immunity. Although inflammation and coagulation are known to have bidirectional relationships, little is known about the interaction between PMN leukocytes and coagulation factors. Coagulation FXI participates in the intrinsic coagulation pathway upon its activation, contributing to hemostasis and thrombosis. We have shown previously that FXI-deficient mice have an increased survival and less leukocyte accumulation into the peritoneum in severe polymicrobial peritonitis. This result suggests a role for FXI in leukocyte trafficking and/or function. In this study, we characterized the functional consequences of FXIa binding to PMN leukocytes. FXIa reduced PMN leukocyte chemotaxis triggered by the chemokine, IL-8, or the bacterial-derived peptide, fMLP, perhaps as a result of the loss of directed migration. In summary, our data suggest that FXIa modulates the inflammatory response of PMN leukocytes by altering migration. These studies highlight the interplay between inflammation and coagulation and suggest that FXIa may play a role in innate immunity.     

Effect of a leukotriene B4 receptor antagonist on leukotriene B4-induced neutrophil chemotaxis in cavine dermis

Leukotriene B₄ (LTB₄) is a proinflammatory product of arachidonic acid metabolism that has been implicated as a mediator in a number of inflammatory diseases. When injected intradermally into the cavine, LTB₄ elicits a dose-dependent immigration (chemotaxis) of neutrophils (PMNs) into the injection sites as assessed by the presence of a neutrophil marker enzyme myeloperoxidase. SC-41930 {7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)propoxy]-3,4-dihydro-8-propyl-2H-l-benzopyran-2-carboxylic acid, a potent LTB₄ receptor antagonist inhibited the chemotactic actions of LTB₄ when coadministered into the dermal site and when given intravenously or orally with ED₅₀ values of 200 ng, 0.5 mg/kg, and 0.6 mg/ kg respectively. This compound may well have application in disease states, such as inflammatory bowel disease and psoriasis, where LTB₄ is implicated as a proinflammatory mediator.     

Orally administered tolfenamic acid inhibits leukotriene synthesis in isolated human peripheral polymorphonuclear leukocytes

Special interest has been focused on the development of dual inhibitors of the cyclo-oxygenase and lipoxygenase pathways of arachidonic acid metabolism. In contrast to other classic NSAIDs, some fenamates in clinically achievable concentrations have been shown to inhibit synthesis of 5-lipoxygenase productsin vitro. In the present work, we studied the effect of orally administered tolfenamic acid (600 mg) on Ca ionophore A 23187-induced leukotriene synthesis in isolated human polymorphonuclear leukocytes. Leukotriene production was reduced in all 14 subjects studied, the mean inhibition of LTB₄ synthesis being 16±3% and that of LTC₄ 33±7%. The inhibition correlated positively with serum tolfenamic acid concentrations. We suggest that inhibition of leukotriene synthesis is an additional mechanism of the anti-inflammatory, antimigraine and antidysmenorrhoeic effects of tolfenamic acid, and a possible explanation for its rare gastric and bronchoconstrictive side-effects.     

Human polymorphonuclear leukocytes release leukotriene B4 during phagocytosis ofStaphylococcus aureus

We studied release of leukotriene B₄ (LTB₄) by human polymorphonuclear leukocytes (PMNs) during phagocytosis of staphylococci in the presence or absence of arachidonic acid. The 12×10⁷ PMNs incubated with 3×10⁹ opsonizedS. aureus and 50M arachidonic acid released 1.45±0.42 nmol LTB₄. No LTB₄ was detected after stimulation of PMNs withS. aureus or arachidonic acid by themselves. However, by increasing the concentration of arachidonic acid to 200 or 400M, 1.22±0.45 and 1.98±0.49 nmol LTB₄, respectively, was released by PMNs. The effect of different bacteria-PMN ratios on LTB₄ production was also studied. LTB₄ varied from 0.3 to 2.0 nmol when bacteria/PMN ratios increased from 5 to 50 (respectively) in the presence of 50 M arachidonic acid. Thus, phagocytizing PMNs produce LTB₄ in the presence of arachidonic acid, and its production is dependent on the number of bacteria phagocytized.     

Leukotriene B4 modulates phospholipid methylation and chemotaxis in human polymorphonuclear leukocytes

Formation of phosphatidylcholine from phosphatidylethanolamine via the S-adenosylmethionine (AdoMet) pathway has been shown to be required for signal transduction of receptor-ligand interactions in a variety of cells. These interactions result in the remodeling of phospholipid pools and phospholipase activation. To extend these observations and to explore the role of the phosphatidylcholine synthesis pathway in transduction of the leukotriene B4 (LTB4) receptor-ligand response, we examined phospholipid methylation in human polymorphonuclear leukocytes (PMN) following stimulation by LTB4, a potent chemotactic agent that is a metabolite of arachidonic acid. At early time points (approximately 3-10 min), formation of methylated phospholipids was enhanced following LTB4 stimulation. The LTB4 analogs 6-trans LTB4 as well as LTB4 epimers induced less methylation compared with LTB4, and the potencies of these analogs in inducing methylation correlated with their diminished ability to induce chemotaxis. Furthermore, the ability of these agonists to induce methylation also correlated with the binding affinity of these agents to the LTB4 receptors on these cells. Synthesis of phosphatidylcholine by the choline transferase pathway was not affected by LTB4. Inhibition of the AdoMet reaction with 3- deazaadenosine, L-homocysteine homolactone, or erythro-9-[2-hydroxy-3-nonyl] adenine (EHNA) abrogated LTB4-induced phospholipid methylation and the chemotactic response. The potencies of these inhibitors in blocking phospholipid methylation also correlated with their ability to abrogate the LTB4-induced chemotactic response. These data suggest that phospholipid methylation and phospholipase activation play an important role in transduction of the LTB4 receptor-ligand interaction in PMN, which results in chemotaxis.     

Pasteurella haemolytica leukotoxin enhances production of leukotriene B4 and 5-hydroxyeicosatetraenoic acid by bovine polymorphonuclear leukocytes.

The influence of the leukotoxin of Pasteurella haemolytica on the generation of arachidonic acid metabolites by bovine polymorphonuclear leukocytes (PMNs) was investigated. PMNs released 5-, 12-, and 15-hydroxyeicosatetraenoic acids (5-, 12-, and 15-HETE) and leukotriene B4 (LTB4) upon stimulation with arachidonic acid. The leukotoxin preparations dose dependently enhanced the release of the 5-lipoxygenase products 5-HETE and LTB4 in arachidonic acid-stimulated PMNs, whereas the release of 12- and 15-HETE was not affected. The enhanced release of LTB4 and 5-HETE was not due to a decreased cellular retention of the 5-lipoxygenase products. In addition, leukotoxin preparations by themselves were also able to induce LTB4 and 5-HETE production in the absence of exogenous arachidonic acid. Generation of 5-lipoxygenase products by PMNs stimulated by leukotoxin may represent an important cellular event that occurs during infections with P. haemolytica.     

Interactions between synthesis of platelet-activating factor and leukotriene B4 in isolated human polymorphonuclear leukocytes

The aim of the present work was to study interactions between the synthesis of platelet-activating factor (PAF) and leukotriene B₄ (LTB₄) in human polymorphonuclear leukocytes (PMNs) in vitro. PAF, at nanomolar concentrations, stimulated calcium ionophore A23187-activated PMNs to release LTB₄ and 5-hydroxyeicosatetraenoic acid (5-HETE). This seems to be a receptor-mediated process as it was blocked by a PAF receptor antagonist WEB 2086 (IC₅₀ 6.6±3.9M). Moreover, LTB₄ stimulated the formation of PAF in activated PMNs. WEB 2086 did not, however, alter PMN migration towards either LTB₄ or the chemotactic peptide FMLP. This suggests that the enhancement of PAF synthesis in response to LTB₄ is a concomitant event rather than a mediating process in LTB₄-induced chemotactic movement of PMNs. These effects are implicated in the complex network of interactions between inflammatory mediators that results accumulation and activation of PMNs in the exacerbation of inflammatory processes.     

Down-regulation of receptor antigen in leukotriene B4-induced chemotactic deactivation of human polymorphonuclear leucocytes.

Pretreatment of suspensions of human polymorphonuclear leucocytes (PMNL) with leukotriene B4 (LTB4) induces chemotactic deactivation, characterized by diminished expression of high-affinity LTB4 receptors and selectively decreased chemotactic responsiveness of the PMNL to LTB4. Rabbit anti-idiotypic antibodies (a-Id) to mouse monoclonal anti-LTB4, which bind to 60,000-80,000 molecular weight (MW) membrane protein of PMNL receptors for LTB4 in Western blots and block binding of [3H]LTB4 to high-affinity receptors of PMNL, detected a reduction in LTB4 receptor antigen during chemotactic deactivation. Loss of high-affinity receptors for LTB4 from the surface of PMNL deactivated by incubation with 10 nM LTB4 was significant after 1 min and after 20 min reached a mean maximum of 82% and 61%, respectively, as assessed by binding of [3H]LTB4 and a-Id. Inhibitors of PMNL proteases did not prevent the deactivation-induced decreases in surface receptors for LTB4. Disruption of deactivated PMNL and solubilization of membrane proteins failed to expose intracellular LTB4 receptors. Incubation of membranes isolated from PMNL with 100 nM LTB4 resulted in a loss of LTB4 receptors similar to that observed in intact PMNL. Changes in LTB4 receptor protein structure or membrane localization, rather than endocytosis or proteolysis, thus appear to explain the rapidly decreased expression of LTB4 receptors, which results from stimulus-specific deactivation.     

Galectin-3 inhibits the chemotaxis of human polymorphonuclear neutrophils in vitro

In the recent years, the participation of the animal lectin galectin (gal)-3 in inflammation and in host defence mechanisms was extensively studied. In vivo studies implied - among others - a role of gal-3 in the recruitment of polymorphonuclear neutrophils (PMN) to sites of bacterial infection. In that context, we asked the question whether gal-3 was chemotactic for PMN. Functional assays revealed that gal-3 was not chemotactic for PMN, but that it inhibited the spontaneous migration and the chemotaxis of PMN towards complement C5a, interleukin (IL)-8, or ATP. Moreover, gal-3 inhibited the shape change and the actin polymerisation of PMN that occurs in response to C5a or IL-8. By use of FITC-labelled gal-3, we found that it attached rapidly to the PMN membrane in a lactose-sensitive manner. In response to gal-3 the MAP kinase p38 was phosphorylated. This kinase is crucial for the migration of PMN towards end-target chemokines, such as C5a, and is activated in response to C5a or IL-8. When PMN were preincubated with gal-3, the C5a-induced p38 phosphorylation was transiently enhanced, but eventually down-modulated. We conclude that by interfering with the chemokine-induced p38 phosphorylation gal-3 inhibits chemotaxis of PMN.     

Decreased release of leukotriene B4 from monocytes and polymorphonuclear leukocytes in patients with systemic lupus erythematosus

The leukotriene B4 (LTB4) releasing capacities of monocytes and polymorphonuclear leukocytes (PMN) were studied using a specific radioimmunoassay in 28 patients with systemic lupus erythematosus (SLE) and 9 normal controls. LTB4 release from both monocytes (7.3 +/- 3.3 ng) and PMN (6.6 +/- 3.4 ng) in SLE patients was decreased compared with that (15.2 +/- 4.3 ng for monocytes, 20.7 +/- 4.5 ng for PMN) in normal controls. There were no differences in the releasing capacities of monocytes and PMN between patients with active and inactive disease. LTB4 suppressed lymphocyte blastogenesis by PHA-P and induced suppressor cells. The significance of the decreased release of LTB4 from monocytes and PMN in SLE patients was discussed.     

Feline polymorphonuclear leukocytes respond chemotactically to leukotriene B4 and activated serum but not to F-met-leu-phe

The chemotactic response of feline polymorphonuclear leukocytes (PMNs) to three types of chemoattractants was studied. Feline PMNs responded to leukotriene B₄ as well as to agarose-activated autologous and homologous serum. However, no response was obtained to N-formylmethionylleucylphenylalanine (FMLP), and four similar peptides that activate the FMLP receptor (N-formylnorleucylleucylphenylalanine, N-formylmethionylphenylalanine, methionylleucylphenylalanine, and pepstatin.) Thus, feline PMNs are similar to equine, porcine, bovine and canine PMNs which also do not respond chemotactically to these peptides.     

Effects of antirheumatic drugs on leukotriene B4 and prostanoid synthesis in human polymorphonuclear leukocytesin vitro

The effects ofd-penicillamine, sodium aurothiomalate, indomethacin, timegadine and tolfenamic acid on the lipoxygenase and cyclo-oxygenase pathways of arachidonic acid metabolism were studied in human polymorphonuclear leukocytes (PMNs)in vitro. In short-term incubations,d-penicillamine and aurothiomalate did not affect leukotriene B₄ (LTB₄), prostaglandin E₂ (PGE₂) or thromboxane B₂ (TXB₂) production. Each of the three non-steroidal anti-inflammatory drugs (NSAIDs) used were potent inhibitors of prostanoid synthesis. In higher concentrations they also reduced LTB₄ production; timegadine and tolfenamic acid were effective in concentrations comparable to those measured in plasma during drug therapy, whereas indomethacin was needed in ten times higher concentrations. The different effects of NSAIDs on 5-lipoxygenase activity may be of importance in their therapeutic actions as well as in the appearance of some side-effects, e.g. gastric irritation and aspirin-induced asthma.     

缺氧对大鼠中性粒细胞趋化能力的影响

目的：研究急、慢性缺氧对大鼠中性粒细胞（PMNs）趋化能力的影响。方法:应用细胞微管吸吮实验技术，从单细胞角度研究在血清趋化作用下不同缺氧状态PMNs伪足生长的变化。结果:自身血清作趋化剂，急性、慢性缺氧组PMNs伪足长度分别为(1.200±0.178)μm和(1.094±0.164)μm，显著大于常氧对照组(0.914±0.156)μm（P＜0.05）；常氧、急性缺氧和慢性缺氧血清作用于常氧PMNs时，急、慢性缺氧血清作用下PMNs伪足长度分别为(1.059±0.179)μm和(1.041±0.175)μm，显著大于常氧血清作用（P＜0.05）；常氧血清作用于常氧、急性和慢性缺氧PMNs时，急性、慢性缺氧PMNs伪足长度分别为(1.058±0.163)μm和(1.118±0.126)μm，显著大于常氧对照组（P＜0.05）。结论:急、慢性缺氧促进了大鼠PMNs趋化能力，缺氧后血液微环境的变化和PMNs响应能力的变化是其重要原因。