Effects of non-steroidal anti-inflammatory drugs on polymorphonuclear leukocyte functions in vitro: focus on fenamates

Prostanoid-independent anti-rheumatic effects of non-steroidal anti-inflammatory drugs (NSAIDs) are a matter of debate. The aim of the present study was to compare the effects of chemically different NSAIDs (diclofenac, indomethacin, ketoprofen, paracetamol, piroxicam and four fenamates: flufenamic, meclofenamic, mefenamic and tolfenamic acids) on human polymorphonuclear leukocyte (PMN) functions, i.e. calcium ionophore A23187-triggered degranulation, leukotriene 134 (LTB4) release, platelet-activating factor (PAF) production and migration towards LTB4. The four fenamates caused a dose-dependent inhibition of each of the PMN functions tested. Flufenamic, meclofenamic and tolfenamic acids were about equipotent to inhibit PMN degranulation (IC₅₀s 21–32 M) and LTB4 release (IC₅₀s 21–25 M) whereas mefenamic acid achieved similar effects at somewhat higher drug concentrations. Tolfenamic and meclofenamic acids were the most potent fenamates to inhibit PAF synthesis (IC₅₀s 37 and 51 M) as well as migration towards LTB4 (IC₅₀s 61 and 92 M). Out of the other NSAIDs, diclofenac (which is chemically related to fenamates) suppressed degranulation as well as LTB4 and PAF production. Indomethacin inhibited LTB₄ and PAF synthesis whereas ketoprofen reduced degranulation. The inhibitory effects of the non-fenamate NSAIDs occurred only at drug concentrations far higher than those achieved clinically. Paracetamol and piroxicam (up to 300 M) did not influence the PMN functions tested. We conclude that NSAIDs with a fenamate structure differ from other NSAIDs by inhibiting PMN functions induced either by receptor-mediated stimulus (LTB4) or calcium ionophore (A23187) at micromolar drug concentrations. Correspondence to: E. Moilanen at the above address     

Effects of the non-steroidal anti-inflammatory drug benoxaprofen on leucocyte migration

Possible modes of action of the anti-inflammatory drug, benoxaprofen, have been explored. The drug caused inhibition of leucocyte migration, principally of mononuclear cells into the pleural cavity of rats undergoing carrageenan-induced pleurisy. Evidence was obtained from in vitro leucocyte migration and chemotaxis models that the drug acted directly on the mononuclear cells rather than by inhibition of chemotactic factors.     

Effects of the non-steroidal anti-inflammatory drug benoxaprofen on leucocyte migration.

Possible modes of action of the anti-inflammatory drug, benoxaprofen, have been explored. The drug caused inhibition of leucocyte migration, principally of mononuclear cells into the pleural cavity of rats undergoing carrageenan-induced pleurisy. Evidence was obtained from in vitro leucocyte migration and chemotaxis models that the drug acted directly on the mononuclear cells rather than by inhibition of chemotactic factors.     

Non-steroidal anti-inflammatory drug-copper complex modulation of polymorphonuclear leukocyte migration

These studies were intended to compare the effects of aspirin, 3,5-diisopropysalicylic acid (3,5-DIPS), and indomethacin with those of their copper complexes: Cu(II)2(aspirinate)4, Cu(II)2(3,5-DIPS)4, and Cu(II)2(indomethacinate)4 as well as Cu(II)2(acetate)4 on polymorphonuclear leukocyte (PMNL) random and directional migration, in addition to their anti-inflammatory activities. Experiments were performed both in vivo and in vitro. In vitro modifications of PMNL migration were measured with the Boyden chamber using N-formyl-methionyl-leucyl-phenylalanine (fMLP) as the chemoattractant and in the agarose assay using fMLP and serum chemotactic derivatives of complement as chemoattractants. In vivo anti-inflammatory activities of these compounds were determined after induction of a serum-induced pleurisy in the rat, and measurement of exudate volume and number of exudative cells 4 hr later. Copper complexes of non-steroidal anti-inflammatory drugs (NSAIDs) were found to be more effective in decreasing random migration and chemotaxis of PMNLs than their parent drugs or Cu(II)2(acetate)4 in in vitro studies. Only chemotaxis was found to be reduced significantly for PMNLs obtained from pleuritic rats after in vivo treatment and the order of copper complex effectiveness was: Cu(II)2(indomethacinate)4 greater than Cu(II)2(3,5-DIPS)4 greater than Cu(II)2(aspirinate)4. All doses of Cu(II)2(acetate)4 administered in vivo failed to affect chemotactic activity. Copper complexes of NSAIDs were also more effective than their parent drugs as anti-inflammatory agents, and Cu(II)2(acetate)4 had no anti-inflammatory activity in this model of inflammation. The order of anti-inflammatory activity was: Cu(II)2(indomethacinate)4 greater than Cu(II)2(3,5-DIPS)4 greater than Cu(II)2(aspirinate)4.     

Effects of non-steroidal anti-inflammatory drugs on lymphocyte activation

Guinea-pig lymph node lymphocytes were stimulated with mitogen (phytohaemagglutinin) in vitro and lymphocyte activation was measured by tritiated thymidine incorporation (DNA synthesis). Inclusion of non-steroidal anti-inflammatory drugs (NSAIDs) in the culture medium at therapeutic concentrations, frequently exerted an inhibitory effect. Such inhibition could not be attributed to the ability of these drugs to inhibit cyclo-oxygenase or lipoxygenase enzymes. Inhibition by salicylates was not associated with cytotoxic or cytopathic effects, since inhibition was only evident when the drugs were included in the early phase of culture. Other NSAIDs exhibited varying degrees of toxicity, which in some instances may account for observed inhibition. The effects on lymphocyte activation of selective inhibitors of pathways of arachidonic acid metabolism, do not support the proposition that the generation of prostaglandins, thromboxanes, leukotrienes or related compounds is an obligatory step during lymphocyte activation.     

Effects of some non-steroidal anti-inflammatory drugs and other agents on cyclooxygenase and lipoxygenase activities in some enzyme preparations

The inhibiting influences of some non-steroidal anti-inflammatory drugs (NSAIDs) and other agents on cyclooxygenase (CO) and lipoxygenase (LO) activities in some enzyme preparations were investigated. Washed rat platelets (CO and 12-LO), rat polymorphonuclear leukocytes (PMNs, CO and 5-LO), rat renal medulla homogenate (CO), and purified soybean LO (15-LO) were used as enzyme preparations. The IC50 values of drugs on the enzyme activities were determined in each preparation. In addition, the inhibitory activities of the drugs on the generation of chemiluminescence from PMNs stimulated by phorbol myristate acetate were tested. NSAIDs (indomethacin, ketoprofen and phenylbutazone) showed a selective inhibition of CO in each preparation, but benoxaprofen inhibited both enzymes, especially in PMNs. BW755C, 1-phenyl-3-pyrazolidone (phenidone), toluene-3,4-dithiol (dithiol), 6-ethoxy-1,2-dihydro-2,2,4-trimethylquinoline (ethoxyquin) and acetone phenylhydrazone (APH) inhibited both enzyme activities. Nordihydroguaiaretic acid (NDGA) showed a relatively selective inhibition of LO in all the preparations used. APH inhibited soybean 15-LO markedly compared with the other enzymes tested. Ethoxyquin inhibited COs more markedly than LOs, and with regard to LO inhibition, it inhibited 5-LO in PMNs markedly. BW755C, phenidone, ethoxyquin, NDGA, APH and dithiol, which strongly inhibited 5-LO, showed an inhibitory activity on the generation of chemiluminescence from PMNs activated with phorbol ester.     

The mode of anti-inflammatory action of a topical non-steroidal anti-inflammatory drug, etofenamate

In order to ascertain the mode of anti-inflammatory action of a topical non-steroidal anti-inflammatory drug, etofenamate which is a diethylene glycol ester of flufenamic acid, the in vitro test for the mechanism of the action were carried out. Etofenamate (3 microM) was hydrolysed to flufenamic acid at a rate of 39.5% and 57.0% of the dose during 30 and 60 min incubation, respectively, when incubated with rat peritoneal macrophages stimulated with starch and bacto peptone in phosphate-buffered saline. PGE2 generation by these cells in MEM medium was dose-relatedly inhibited with etofenamate as well as flufenamic acid at the dosage range of 1 to 30 microM. This suggests that unchanged etofenamate is active, since the highest conversion rate of etofenamate to flufenamic acid was 15% of the dose during the incubation. Etofenamate produced a dose-related inhibition against lipoxygenase prepared from peritoneal polymorphonuclear leucocytes of guinea pigs, and its activity (IC50 = 5.3 X 10(-5) M) was stronger than that of caffeic acid; flufenamic acid was inactive. Inhibitory activity of etofenamate was one-third or less that of flufenamic acid against the hypotonic-hyperthermic lysis of rat erythrocytes and heat-denaturation of bovine serum albumin. From these results, it was suggested that topically applied etofenamate produces its anti-inflammatory action through prostaglandin synthesis inhibition by flufenamic acid produced in the inflammatory tissue and inhibition of prostaglandin synthesis by macrophages and lipoxygenase inhibition by unchanged etofenamate.     

Inhibition of some polymorphonuclear leukocyte functions by ethacrynic acid

Ethacrynic acid (10(-4) M) inhibits exocytosis, phagocytosis and superoxide release in rabbit polymorphonuclear leukocytes (PMN's). Dihydroethacrynic acid is a much weaker inhibitor of these PMN functions. Though ethacrynic acid inhibits ATPase activity in the PMN, this occur at much higher concentrations than required for inhibition of exocytosis and superoxide release, thus a causal relationship seems unlikely. The same applies to inhibition of ATP generation by ethacrynic acid: the concentration required to decrease ATP level in PMN's is much higher than required for the inhibitory effect on exocytosis. Inhibition of exocytosis by ethacrynic acid can be prevented by dithiothreitol. It is concluded that vulnerable sulfhydryl groups are involved in the inhibition by ethacrynic acid.     

Effects of isoxicam and other non-steroidal anti-inflammatory drugs on arachidonic acid metabolism by rat peritoneal leucocytes

Inhibition of prostaglandin formation from [14C]arachidonic acid by rat peritoneal leucocytes occurred with nonsteroidal anti-inflammatory drugs, their order of potency being indomethacin greater than piroxicam greater than naproxen greater than ibuprofen greater than isoxicam. At the lowest concentration tested (1 microgram ml-1), indomethacin markedly increased the accumulation of lipoxygenase products in the cell incubates. Naproxen, ibuprofen or piroxicam 1 or 10 micrograms ml-1 resulted in smaller increases of lipoxygenase products, and there was only a small rise with these concentrations of isoxicam.     

In vitro effects of non-steroidal anti-inflammatory drugs on human polymorphonuclear cells and lymphocyte migration.

1. The action of the non-steroidal anti-inflammatory drugs (NSAID), sodium salicylate, aspirin, phenylbutazone and indomethacin was investigated on the migration of human polymorphonuclear cells (PMNs) and lymphocytes, using the system of migration of leucocytes from glass capillary tubes. 2. All NSAID produced a dose-dependent inhibition of cell migration, and were more effective on the migration of the PMN than on lymphocytes. 3. Drugs optimally suppressed PMN migration after 20 to 24 h incubation, and lymphocytes after 3 to 6 h. 4. Prolonged incubation of cells with several concentrations of NSAID demonstrated an 'escape' from inhibition in PMNs prepared from one subject.     

非甾体抗炎药致重症药疹文献汇总分析

目的：以临床实例出发探讨非甾体抗炎药致重症药疹的特点和关联性,为临床用药安全提供参考。方法：通过检索1970-2017年国内外期刊数据库公开报道的非甾体抗炎药致重症药疹的病例,提取文献中患者年龄、性别、致ADR药物、重症药疹类型等信息进行统计和分析。结果：非甾体抗炎药致重症药疹文献49篇,共计病例49例。其中单一用药致重症药疹病例有27例,联合用药22例;49例非甾体抗炎药致重症药疹中发生率最高为对乙酰氨基酚,其次为吡罗昔康、布洛芬、依托考昔;非甾体抗炎药致重症药疹平均潜伏期为（7.88±10.42）d,其中致药物超敏反应综合征潜伏期最长,可长达（36.50±14.20）d;大部分患者停药后好转,1例因中毒性表皮坏死松解征死亡。结论：非甾体抗炎药致重症药疹具有潜在的危险性,临床应用时应提高警惕,以减少重症药疹给患者带来的危害。     

Antimutagenic and anti-oxidant activities of the non-steroidal anti-inflammatory drug celecoxib

1. Tumors arise and progress through the accumulation of serial genetic changes, including successive mutations, which involve activation of proto-oncogenes and inactivation of tumour suppressor genes, leading to the uncontrolled proliferation of progeny cells. The human body is continuously and unavoidably exposed to structurally diverse chemicals with established carcinogenic activity in animal models and/or mutagenic activity in short-term tests. 2. Celecoxib, a non-steroidal anti-inflammatory drug that specifically inhibits the enzyme cyclo-oxygenase-2, has been reported to be effective against certain types of cancers. The in vitro anti-oxidant and antimutagenic activities of the celecoxib were investigated in the present study using standard procedures. 3. The antimutagenic activity of celecoxib was determined using histidine mutant Salmonella typhimurium strains TA98, TA100, TA102 and TA1535 against directly acting mutagens (sodium azide (NaN3), N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), 4-nitro-o-phenylenediamine (NPDA) and doxorubicin) and mutagens needing activation (2-acetamidofluorene (2-AF) and 7,12-dimethylbenz [a] anthracene (DMBA)). 4. Celecoxib inhibited NaN3-, MNNG- and NPDA-induced mutations of TA100. The antimutagenicity of celecoxib (0.2 mg/plate) against the NaN3-induced mutation of TA1535 was 39.8% (P < 0.001). The MNNG-induced mutation of TA1535 was also inhibited by 0.3 mg/plate celecoxib (46.0%; P < 0.05). At concentrations of 0.2 mg/plate, celecoxib significantly inhibited NPDA- and doxorubicin-induced mutations of TA98 by 52.5 and 58.0%, respectively (P < 0.001 and P < 0.05, respectively). 5. The antimutagenic activity of 0.3 mg/plate celecoxib against 2-AF- and DMBA-induced mutations of TA98 was 81.76 and 98.1%, respectively (P < 0.001). 6. The anti-oxidant activity of celecoxib was determined by the inhibition of lipid peroxidation and superoxide and hydroxyl radical-scavenging activities. 7. The IC50 values of celecoxib for hydroxyl radical-scavenging and the inhibition of lipid peroxidation were 1.97 +/- 0.06 and 1.99 +/- 0.05 micromol/mL, respectively. Celecoxib had no superoxide radical scavenging-activity up to a concentration of 2.6 micromol/mL. 8. The in vitro antimutagenic and anti-oxidant activities of celecoxib indicate its possible therapeutic use as a cancer chemopreventive agent.     

On the mechanism of the anti-inflammatory activity of some copper (II) complexes

The mechanism of the antiinflammatory activity of three copper(II) complexes: tetrachlorocuprate(II) of bis(2.4.6-trimethylpyridinio), tetrachlorocuprate(II) of bis(2.4-dimethylpyridinio) and tetrachlorocuprate(II) of bis-pyridinio, has been investigated. The antiinflammatory activity does not seem to be related to an inhibitory effect on prostaglandin biosynthesis or with leukocyte migration into inflamed tissue. These complexes showed an inhibitory effect on volume exudate in experimental pleurisy. They showed also a potent inhibitory effect on the production of superoxide anions. These inhibitory effects could explain, at least in part, their antiinflammatory activity.     

Pharmacological properties of a new non-steroidal anti-inflammatory drug: flunoxaprofen

The anti-inflammatory, analgesic and antipyretic activities of S-(+)-2(4-fluorophenyl)-alpha-methyl-5 benzoxazole acetic acid (flunoxaprofen: Flu), a new non-steroidal anti-inflammatory drug, were compared with those of indomethacin and other non-steroidal anti-inflammatory drugs (NSAIDs) in experimental animals. Flu showed strong inhibitory activity on acute and subacute inflammation tests in rats, such as carrageenin hind paw oedema (oral: 6-25 mg/kg; rectal: 50-100 mg/kg); pellet-induced granuloma formation (5-20 mg/kg/day) and adjuvant-induced arthritis (10 mg/kg/day). Its potency was comparable with that of indomethacin (I) and higher than that of acetyl salicylic acid (ASA), ibuprofen (IBU) or phenylbutazone (P). The analgesic activity of Flu, evaluated by the hot plate method and tail pinching in mice, was slightly lower than that of I but higher than that of ASA and IBU. In pyretic rabbits Flu showed an antipyretic activity higher than that of ASA and IBU. The ability of Flu to affect platelet aggregation, mucopolysaccharide synthesis by fibroblasts and the proteolytic action of trypsin was also investigated.     

Inhibition of peroxidase activity by some non-steroidal anti-inflammatory drugs.

The non-steroidal anti-inflammatory drugs, indomethacin, flufenamic acid and naproxen inhibited thyroid peroxidase-catalyzed iodination of BSA in vitro. Inhibition by all three drugs was affected more effectively in a hydrogen peroxide generating system than in an incubation system in which hydrogen peroxide was added. Naproxen differed from the other two drugs in so far as it inhibited mainly hydrogen peroxide generation while its comparatively low inhibitory influence (>20%) on TPO was not dose-dependent. The inhibitory influence of these anti-inflammatory drugs was also observed when other peroxidases, such as lactoperoxidase, chloroperoxidase and horseradish peroxidase were used for catalyzing BSA iodination in a hydrogen peroxide generating system. No iodination of BSA was obtained with horseradish peroxidase when hydrogen peroxide was added instead of generated so that the inhibitory nature of these drugs could not distinguish between their direct effect on horseradish peroxidase or on hydrogen peroxide generation. However, in lactoperoxidase and chloroperoxidase-catalyzed BSA iodinations in non-H₂O₂-generating systems naproxen had no appreciable inhibitory influence below a 1 millimolar concentration. On another thyroid peroxidase activity, namely its catalyzing influence on the exchange reaction between inorganic iodide and organic iodine in diiodotyrosine, indomethacin and naproxen showed unappreciable effects lower than 1 mM concentrations. Similarly, on thyroid peroxidase and lactoperoxidasecatalyzed deiodination of thyroxine they were ineffective inhibitors. The data suggest that these antiinflammatory drugs are effective inhibitors of iodination reactions but ineffective inhibitors of deiodination reactions.