Prostaglandin biosynthesis in rabbit kidney medulla: inhibition in-vitro vs. in-vivo by aspirin, indomethacin and meclofenamic acid.

The non-steroidal anti-inflammatory drugs aspirin, indomethacin and meclofenamic acid were compared for their potency and duration of inhibition of prostaglandin biosynthesis in rabbit kidney medulla. Indomethacin and meclofenamic acid showed equal potency of inhibition in-vitro (IC50 0.88 micron and 0.85 micron respectively) while aspiring was a much weaker inhibitor (IC50 120 micron). In-vivo, indomethacin was the most powerful inhibitor (ID50 0.034 mg/kg) followed by meclofenamic acid (0.45 mg/kg) and aspirin (2.35 mg/kg). Studies on the duration of in-vivo inhibition by these compounds showed the effect of indomethacin and meclofenamic acid to be completely reversed within 4-6 hours. In contrast, return of kidney prostaglandin biosynthetic activity following aspirin inhibition is very slow and significant inhibition is still present 48 hours after a single aspiring injection. The inhibitory effect of aspirin in-vivo could be blocked by pretreatment with indomethacin, indicating that both drugs interact with related sites on the cyclo-oxygenase enzyme. The irreversible inhibition of the cyclo-oxygenase by aspirin as demonstrated in studies of other investigators suggests that the return of kidney prostaglandin synthetase activity after aspirin inhibition represents synthesis of new cyclo-oxygenase protein.     

Fenoprofen: Inhibitor of prostaglandin synthesis

Fenoprofen, an anti-inflammatory agent, is a potent inhibitor of prostaglandin synthesis at physiological substrate concentration. I₅₀'s of 1 μM to 100 μM were obtained when the concentration of arachidonate was increased from 1 μM to 82 μM. In contrast, a small change in I₅₀ was observed with indomethacin. Kinetic studies indicated that inhibition by fenoprofen was competitive while that of indomethacin was non-competitive.     

Inhibition of prostaglandin synthesis in mouse 3T3 fibroblasts and human platelets by substituted phenols

Several substituted phenols with antioxidant properties were potent reversible inhibitors of prostaglandin synthesis in 3T3 cell cultures. The ID₅₀'s for prostaglandin (PG) E₂ synthesis in these cells were 0.1 μM for 2,6-xylenol, 5 μM for tricresol, 6 μM for -cresol, 7 μM for -cresol, 15 μM for 3,5-xylenol, 30 μM for -cresol and 100 μM for phenol. The corresponding values for aspirin and indomethacin were 4 μM and 0.02 μM, respectively.The substituted phenols also inhibited serotonin release, aggregation and prostaglandin synthesis in human platelets induced by arachidonic acid but not by PGG₂.     

Synthesis and biological evaluation of 1-(benzenesulfonamido)-2-[5-(N-hydroxypyridin-2(1H)-one)]acetylene regioisomers: a novel class of 5-lipoxygenase inhibitors

A hitherto unknown class of linear acetylene regioisomers were designed such that a SO₂NH₂ group was located at the ortho-, meta-, or para-position of the acetylene C-1 phenyl ring, and a N-hydroxypyridin-2(1H)-one moiety was attached via its C-5 position to the C-2 position on an acetylene template (scaffold). All three regioisomers inhibited 5-lipoxygenase (5-LOX), where the relative potency order was 2-SO₂NH₂ (IC₅₀ = 10 μM) >3-SO₂NH₂ (IC₅₀ = 15 μM) >4-SO₂NH₂ (IC₅₀ = 68 μM) relative to the reference drug nordihydroguaiaretic acid (NDGA; IC₅₀ = 35 μM). The 2-SO₂NH₂ regioisomer (ED₅₀ = 86.0 mg/kg po) exhibited excellent oral anti-inflammatory (AI) activity that was more potent than aspirin (ED₅₀ = 128.9 mg/kg) and marginally less potent than ibuprofen (ED₅₀ = 67.4 mg/kg). The N-hydroxypyridin-2(1H)one moiety provides a novel pharmacophore for the design of cyclic hydroxamic mimetics capable of chelating 5-LOX iron for exploitation in the design of 5-LOX inhibitory AI drugs.     

Antagonism by ETYA of the effects of leukotrienes on ileum and lung parenchymal strips independent of effects on arachidonic acid metabolism

5,8,11,14-Eicosatetraynoic acid (ETYA), a compound which inhibits both the cyclooxygenase and lipoxygenase pathways of arachidonic acid metabolism, antagonized the contraction of segments of guinea-pig ileal longitudinal muscle produced by SRS-A (IC₅₀ = 2.73 μM). This activity was unaffected by pretreatment of the tissues with 10 μM indomethacin. Phenidone, another mixed cyclooxgenese-lipoxygenese inhibitor, was inactive. FPL-55712, an SRS-A antagonist, was a very potent inhibitor (IC₅₀ = 0.011 μM).BW755C and NDGA nonselectively inhibited the contractions of the guinea-pig ileal longitudinal muscle induced by SRS-A or histamine.ETYA antagonized the contraction of the guinea-pig ileal strip produced by 6 nM synthetic LTC₄ (IC₅₀ = 9.3 μM). FPL-55712 demonstrated an IC₅₀ of 0.3 μM in a similar series of experiments. ETYA, 1, 3 or 10 μM did not inhibit the contractions elicited by 0.5 μM of histamine.This was not a tissue-selective effect since 100 μM ETYA antagonized the LTC₄-induced contraction of the guinea-pig lung parenchymal strip preparation.These data demonstrate that ETYA antagonized the contractile effect of the leukotrienes on tissues from the gastrointestinal tract and lung. Furthermore, the inability of indomethacin or phenidone to inhibit the contractile response suggests that antagonism by ETYA may occur by a mechanism independent of cyclooxygenase and lipoxygenase enzymes.     

Food Polyphenols Fail to Cause a Biologically Relevant Reduction of COX-2 Activity

Epidemiologic studies show a correlation between the dietary intake of food polyphenols and beneficial health effects. Several in vitro studies indicate that the anti-inflammatory potential of polyphenols is, at least in part, mediated by a modulation of the enzymes of the arachidonic acid cascade, such as the prostaglandin forming cyclooxygenases (COXs). Evidence that this mode of action can be transferred to the situation in vivo is scarce. This study characterized effects of a subset of polyphenols on COX–2 expression and activity in vitro and compared the potency with known drugs. Next, the in vivo relevance of the observed in vitro effects was tested. Enzyme assays and incubations of polyphenols with the cancer cell line HCA–7 and lipopolysaccharide (LPS) stimulated primary monocytes support the hypothesis that polyphenols can effect COX–2 expression and activity in vitro. The effects were most pronounced in the monocyte assay for wogonin, apigenin, resveratrol and genistein with IC₅₀ values of 1.5 μM, 2.6 μM, 2.8 μM and 7.4 μM. However, these values are 100- to 1000-fold higher in comparison to those of the known pharmaceuticals celecoxib, indomethacin and dexamethasone. In an animal model of LPS induced sepsis, pretreatment with polyphenols (i. p. 100 mg/kg bw) did not result in decreased plasma or tissue prostaglandin levels, whereas the positive control celecoxib effectively attenuated LPS induced prostaglandin formation. These data suggest that despite the moderate potency in vitro, an effect of polyphenols on COX–2 during acute inflammation is unlikely, even if a high dose of polyphenols is ingested.     

Naturally occurring conjugated octadecatrienoic acids are strong inhibitors of prostaglandin biosynthesis

Fatty acids from natural sources (mostly seed oils) were isolated and assayed for their effect on the bioconversio of arachidonic acid into prostaglandin E₂, using sheep vesicular gland microsomes. Homologues and isomers of the naturally occurring fatty acids, obtained by chemical modification and/or organic synthetic methods, were also tested. Two very active cyclooxygenase inhibitors were discovered, namely jacarandic acid (8 , 10 , 12 -octadecatrienoic acid), isolated from , the concentration which gives 50% inhibition ([I]₅₀) being 2.4 μM and the synthetic 8 , 10 , 12 -octadecarienoic acid, having an [I]₅₀ of 1.0 μM. Under the conditions of the assay (75 μM substrate), earlier described potent inhibitors showed the following [I]₅₀′s: indomethacin: 1.3 μM; 9,12-octadecadiynoic acid: 1.3 μM, 8 , 12 , 14 -eicosatrienoic acid: 2.7 μM; 5,8,11,14-eicosatetraynoic acid: 4.4 μM. At a concentration of about half that of the substrate, the following naturally occurring fatty acids revealed inhibition ([I]₅₀): columbinic acid (29 μM), calendulic acid (31 μM), liagoric acid (31 μM), ximenynic acid (39 μM), crepenynic acid (40 μM) and timnodonic acid (43 μM). Other fatty acids, and some of the above acids, were converted themselves more or less rapidly, mostly into conjugated monohydroxy fatty acids.     

Angiotensin I-converting enzyme inhibitory peptides in red-mold rice made by Monascus purpureus

The ACE inhibitory activity in red-mold rice extracts, prepared from 24 strains of the genus Monascus, was measured. The most effective strain for ACE inhibition was Monascus purpureus IFO 4489 (IC₅₀ = 0.71 mg/ml). Although the antihypertensive substance γ-amino butyric acid was detected in the red-mold rice (85.2 mg/kg), it did not contribute to ACE inhibition. Four ACE inhibitory peptides were isolated from the extract and identified as Ile-Tyr (IC₅₀ = 4.0 μM), Val-Val-Tyr (22.0 μM), Val-Phe (49.7 μM) and Val-Trp (3.1 μM) by protein sequencing. The ACE inhibitory activity of these peptides was almost completely preserved after successive in vitro digestion by pepsin, chymotrypsin and trypsin. These results suggest that red-mold rice made by M. purpureus could be useful in alleviating hypertension.     

The (+)- and (−)-gossypols potently inhibit human and rat 11β-hydroxysteroid dehydrogenase type 2

Gossypol has been proven to be a very effective male contraceptive. However, clinical trials showed that the major side effect of gossypol was hypokalemia. Gossypol occurs naturally as enantiomeric mixtures of (+)-gossypol and (−)-gossypol. The (−)-gossypol is found to be the active component of antifertility. 11β-Hydroxysteroid dehydrogenase 2 (11βHSD2) has been demonstrated to be a mineralocorticoid receptor (MR) protector by inactivating active glucocorticoids including corticosterone (CORT) in rats, and therefore mutation or suppression of 11βHSD2 causes hypokalemia and hypertension. In the present study, the potency of gossypol enantiomers was tested for the inhibition of 11βHSD1 and 2 in rat and human. Both (+) and (−)-gossypols showed a potent inhibition of 11βHSD2 with the half maximal inhibitory concentration (IC₅₀) of 0.61 and 1.33 μM for (+) and (−)-gossypols, respectively in rats and 1.05 and 1.90 μM for (+) and (−)-gossypols, respectively in human. The potency of gossypol to inhibit 11βHSD1 was far less; the IC₅₀ was ≥100 μM for racemic gossypol. The gossypol-induced hypokalemia is likely associated with its potent inhibition of kidney 11βHSD2.     

Effect of the non-steroidal anti-inflammatory drugs on the acyl-CoA synthetase activity toward medium-chain,long-chain and polyunsaturated fatty acids in mitochondria of mouse liver and brain

Effect of eleven non-steroidal anti-inflammatory drugs on the acyl-CoA synthetase activities toward octanoic, palmitic, arachidonic and docosahexaenoic acids was evaluated in mouse liver and brain mitochondria. The drugs tested were aspirin, salicylic acid, diflunisal, mefenamic acid, indomethacin, etodolac, ibuprofen, ketoprofen, naproxen, loxoprofen, flurbiprofen. In mouse liver mitochondria, diflunisal and mefenamic acid exhibited the inhibitory activities not only for octanoic acid (IC₅₀?=?78.7 and 64.7 µM) and but also for palmitic acid (IC₅₀?=?236.5 and 284.4 µM), respectively. Aspirin was an inhibitor for the activation of octanoic acid only (IC₅₀?=?411.0 µM). In the brain, mefenamic acid and diflunisal inhibited strongly palmitoyl-CoA formation (IC₅₀?=?57.3 and 114.0 µM), respectively. The activation of docosahexaenoic acid in brain was sensitive to inhibition by diflunisal and mefenamic acid compared with liver.     

Effect of indomethacin and 7-oxa-13-prostynoic acid on luteinization of transplanted rat ovarian follicles induced by luteinizing hormone and prostaglandin E2

The ability of prostaglandin E₂ (PGE₂) to initiate luteinization was demonstrated using a system of in vitro incubation of ovarian follicles followed by transplantation. Follicles from diestrous rats were incubated with 0.05 to 50 μg/ml PGE₂, 10 μg/ml luteinizing hormone (LH), or alone in Krebs-Ringer bicarbonate buffer plus glucose for 2 hr. Then follicles were transplanted under the kidney capsules of hypophysectomized recipients, with follicles exposed to PGE₂ on one side and those exposed to LH or buffer only on the other side. As determined at autopsy 4 days later and confirmed by histological examination, follicles exposed to PGE₂ at concentrations of 0.5 μg/ml or greater, or to LH, transformed into corpora lutea, but control follicles regressed. Incubation of follicles with LH in the presence of indomethacin, an inhibitor of prostaglandin synthesis, significantly reduced the incidence of luteinization. Prostaglandin E₂ (10 μg/ml) was able to override the inhibition of luteinization by indomethacin (150 μg/ml). The prostaglandin analogue 7-oxa-13-prostynoic acid (100 μg/ml) failed to prevent luteinization in response to either 5 μg/ml LH or 1 μg/ml PGE₂. Results with PGE₂ and with indomethacin suggest a role for prostaglandins in the luteinizing action of LH.We have reported previously that in vitro exposure of diestrous rat follicles to luteinizing hormone (LH) will result in transformation of the follicles to corpora lutea following transplantation under the kidney capsules of hypophysectomized rats. Dibutyryl cyclic AMP (DBC) mimics this effect of LH, and transplants produce progesterone in measurable amounts after both LH and DBC exposure when prolactin is administered in vivo to recipients.Kuehl et al. have suggested that prostaglandins may act as obligatory intermediates in the effect of LH on the ovary, acting between LH and adenylate cyclase. Preliminary results indicated that prostaglandin E₂ (PGE₂) could induce luteinization in our system. The extent of prostaglandin involvement in luteinization was further investigated in this work, using two reported antagonists of prostaglandin action, indomethacin and 7-oxa-13-prostynoic acid. Indomethacin has been found to inhibit synthesis of prostaglandins E₂ and F_2α; 7-oxa-13-prostynoic acid, which acts as a competitive antagonist of prostaglandins, prevented the effect of LH and prostaglandins E₁ and E₂ on cyclic AMP production in mouse ovaries.     

A new prostaglandin E1 analogue (CL-115,574) II. Effects on gastric acid secretion in the dog

The efficacy of CL-115,574, a prostaglandin E₁ analogue, as an acid antisecretory agent was evaluated in dogs. CL-115,574 inhibited acid secretion maximally at an oral dose of 20 μg/kg causing 100% inhibition of acid secretion up to one hour after administration, with significant inhibition of secretion (30%) still present nearly four hours after drug administration. The wide disparity between the maximally effective antisecretory dose 20 μg/kg and the dose at which reproducible side effects occurred (1 mg/kg) suggests that this compound may be developed as an antisecretory compound for use in man.     

Stimulation by melittin of adrenocorticotropin and beta-endorphin release from rat adenohypophysis in vitro

The effect of melittin on the release of adrenocorticotropin (ACTH) and β-endorphin from the corticotropic cells of the rat adenohypophysis was examined in vitro. Anterior pituitary quarters were perifused or incubated in vitro and ACTH- (ACTH-IR) or β-endorphin-like immunoreactivity (β-End-IR) in the medium was measured by radioimmunoassays. Melittin stimulated ACTH-IR and β-End-IR release. This effect was rapid in onset, reversible, and concentration-related (50–5000 ng/ml) and dependend on the presence of calcium ions in the incubation medium. Melittin also elevated the tissue content of unesterified ³H-arachidonic acid that had previously been incorporated into lipids. Purported phospholipase A₂ inhibitors, mepacrine (up to 1 mM), dexamethasone (0.5 mg/kg in vivo, 50 nM in vitro), or p-bromophenacylbromide (100 μM), did not decrease the Melittin (500 ng/ml) — induced β-End-IR release, although mepacrine and dexamethasone may have inhibited phospholipase A₂ activity as indicated by an inhibition of melittin-evoked prostaglandin E₂ formation. After stimulation by melittin (500 ng/ml), β-End-IR release was not affected by the cyclooxygenase inhibitor inddomethacin (up to 140 μM), whereas nordihydroguaiaretic acid (100 μM), a lipoxygenase inhibitor, or BW755C (250 μM), an inhibitor of both cyclooxygenase and lipoxygenase, abolished melittin-induced hormone secretion. We conclude that melittin generates a signal in the corticotropic cells of the rat adenohypophysis which induces hormone secretion by exocytosis. This signal may be unrelated to the activation by melittin of phospholipase A₂.     

Lack of covalent modification of prostaglandin synthetase (cyclo-oxygenase) by indomethacin

We have previously shown that aspirin irreversibly inhibits prostaglandin synthetase (cyclo-oxygenase) by acetylating the active site of the enzyme. By utilizing ¹⁴C-labeled indomethacin and a close analogue, we now show that indomethacin, unlike aspirin, does not covalently modify cyclo-oxygenase. Furthermore, indomethacin binding to the enzyme may be reversible since even though indomethacin can inhibit acetylation by aspirin, when enzyme inhibited by indomethacin (1 μM) is treated with 200 μM aspirin 3 times for 1 hour each, complete acetylation of cyclo-oxygenase is achieved.     

Effects of aspirin and indomethacin on cerebral circulation in the conscious rat: Evidence for a physiological role of endogenous prostaglandins

The purpose of this work was to evaluate the effects of equipotent doses of two different inhibitors of cyclo-oxygenase, indomethacin and aspirin, on cerebral blood flow and cerebral vascular resistances in the conscious undisturbed rat, using the reference sample radioactive microsphere method. We found that both, aspirin (50 mg/kg) and indomethacin (5 mg/kg) at 3, 15 and 60 min after their intravenous administration, increased cerebral vascular resistances and decreased cerebral blood flow to a similar extent. Both drugs completely abolished the hypotensive effect of 5 mg/kg i.v. arachidonic acid and they did not change arterial PO₂, PCO₂ or pH values. We conclude that the pharmacological inhibition of cyclooxygenase in the conscious undisturbed rat leads to a cerebral vasoconstriction and consequently to a decrease in cerebral blood flow. Our results evidence that prostaglandins are a physiological factor that actively contributes to the maintenance of cerebral circulation.     

Lipoxygenase- and cyclooxygenase-reaction products and incorporation into glycerolipids of radiolabeled arachidonic acid in the bovine retina

The metabolism of radiolabeled arachidonic acid (AA) by the intact bovine retina has been studied. Synthesis of prostaglandins (PGs) and hydroxyeicosatetraenoic acids (HETEs), and incorporation of AA into glycerolipids has been measured by reverse-phase and straight-phase high performance liquid chromatography with flow scintillation detection, and by thin-layer chromatography. AA was actively acylated into glycerolipids, particularly triglycerides, phosphatidylcholine and phosphatidylinositol. AA was also converted to the major PGs, PGF_2α, PGE₂, PGD₂, 6-keto-PGF_1α and TXB₂, and to the lipoxygenase reaction products, 12-HETE, 5-HETE, and other monohydroxy isomers. Approximately 6% of the radiolabeled AA was converted to eicosanoids. The synthesis of HETEs was inhibited in a concentration-dependent manner (IC₅₀ = 8.3 NM) by nordihydroguaiaretic acid (NDGA). PG synthesis was inhibited by aspirin (10 μM), indomethacin (1 μM) and NDGA (IC₅₀ = 380 nM). Metabolism of AA via lipoxygenase, cyclooxygenase and activation-acylation was inhibited by boiling retinal tissue prior to incubation. These studies demonstrate an active system for the uptake and utilization of AA in the bovine retina, and provide the first evidence of lipoxygenase-mediated metabolism of AA, resulting in the synthesis of mono-hydroxyeicosatetraenoic acids, in the retina.     

Prostaglandin synthesis inhibited by formamidine pesticides.

The formamidine pesticides, chlordimeform and amitraz, were shown to have both antipyretic and anti-inflammatory activity when given intraperitoneally to rats at 5 to 80 mg per kg. They reduced yeast-induced fever in rats with potencies intermediate between those of indomethacin and aspirin, and antagonized the carageenin-induced swelling of the hind paw. In both these actions, chlordimeform was more potent than amitraz. Both formamidines also inhibited the synthesis of prostaglandin E₂ from arachidonic acid by bovine seminal vesicle microsomes. At an arachidonic acid concentration of 0.4 μM, the I₅₀ values for chlordimeform and amitraz were 34 and 880 μM respectively, compared to 0.4 μM and 790 μM for indomethacin and aspirin. These aspirin-like actions may provide a clue to some of the physiological effects of the formamidines, which represent a new and unsual group of prostaglandin synthetase inhibitors.     

Indomethacin prevents the long-lasting inhibitory effect of aspirin on human platelet cyclo-oxygenase activity

Aspirin and indomethacin do interact with the same site on cyclo-oxygenase. This suggestion is based on studies on ram seminal vesicles and drug interaction studies on rat platelets. The purpose of the present study was to ascertain whether the same interaction also occurred after administration of both drugs to human volunteers.Platelet aggregation induced by sodium arachidonate or by collagen, and formation of platelet MDA and TxB₂ were measured before, two and 48 hours after ingestion of either indomethacin (50 mg) or aspirin (500 mg) or of both drugs (30 minutes apart).While the inhibitory effect of indomethacin on these parameters was short lasting, that of aspirin persisted for at least 48 hours. However, when both drugs were given concurrently, the long-lasting effect of aspirin was no longer detectable. Since competition at levels other than platelets was unlikely, this study indicates that indomethacin and aspirin inhibit human platelet cyclo-oxygenase by the same basic mechanism. Acetylation of the enzyme appears to be a secondary mechanism which makes the inhibitory effect of aspirin persistent.     

Effect of TFC-612, a 7-thia prostaglandin E1 derivative, on a peripheral arterial occlusive disease model in rats

TFC-612, methyl 6-({(1R,2S,3R)-3-hydroxy-2-{(1E,3S,5R)-3-hydroxy-5-methyl-1-nonenyl}-5-oxocyclopentyl}-thio]-hexanoate, inhibited the progression of the lesion in a lauric acid-induced peripheral arterial occlusive model at 1.0 mg/kg p.o. or 1.0 μg/rat/h s.c. in rats. Aspirin (32 mg/kg, p.o.), an anti-platelet drug, did not suppress the lesion growth. On the other hand, ketanserin (10 mg/kg, p.o.), a 5-HT₂ antagonist, also inhibited the progression of the lesion. In vitro, TFC-612 inhibited rat platelet aggregation induced by collagen and ADP with IC₅₀ values of 5.4 ng/mL and 9.5 ng/mL, respectively. Aspirin also inhibited collagen-induced aggregation with an IC₅₀ value of 6.3 μg/mL, but not ADP-induced aggregation at 180 μg/mL. Ketanserin had no effect on either aggregation at 40 μg/mL. In ex vivo experiments, aspirin inhibited platelet aggregation induced by collagen at 10 and 32 mg/kg in rats. However, TFC-612 showed significant inhibition only at 10 mglkg. TFC-612 and ketanserin increased dermal blood flow in the rat paw at 1.0 μg/kg i.v. and 100 μg/kg Lv., respectively. Aspirin had no effect on blood flow at 3.2 mg/kg i.v. These results suggest that the improvement of microcirculation, in addition to anti-platelet action by TFC-612, contributes to its inhibitory effect in a peripheral arterial occlusive model in rats.     

Possible relation of prostaglandins to PMN-derived mediators of host metabolic responses to inflammation

Stimulated rabbit peritoneal polymorphonuclear leukocyte (PMN) preparations simultaneously produce prostaglandin-like material and mediators that induce metabolic alterations in experimental animals characteristic of the host's responses to inflammation. The alterations observed in rats include responses by: proteins, carbohydrates, hormones, trace metals, and total blood neutrophils. This study demonstrates a possible relationship between prostaglandins and PMN-derived substances that mediate plasma zinc depression, hepatic amino acid uptake, and increased numbers of blood neutrophils. Production of these mediators by stimulated-PMN preparations was prevented by 23 μM indomethacin or 93 μM aspirin. Conversely, morphine (2 mM or less) had no detrimental effect on production of these mediators, although, it consistently stimulated production of a substance stimulating total blood neutrophilia. In addition, 2 μM prostaglandin E and F stimulated production of substances mediating hepatic amino acid uptake and plasma zinc depression, respectively. At this concentration, neither prostaglandin significantly altered production of substances mediating increased numbers of total blood neutrophils. A partial-nitrogen atmosphere, dibutyryl cyclic analogs of AMP and GMP, or homogenization of the PMN had no effect on mediator production. The inhibitory effect of indomethacin and aspirin also was observed with PMN-homogenates. These experimental observations suggest that prostaglandin synthesis has a function in production of mediators by stimulated-PMN preparations.