[The pharmacology of the lipoxygenase inhibitors oxphaman (1-(2,5-dihydroxybenzylidene)aminoadamantane) and oxphalin (1-(3,4-dihydroxybenzylidene)-2,4,6-trimethylaniline]. / Zur Pharmakologie der Lipoxygenasehemmer Oxphaman, 1-(2,5-Dihydroxybenzyliden)aminoadamanten, und Oxphalin, 1-(3,4-Dihydroxybenzyliden)-2,4,6-trimethylanilin.

Oxphaman and particularly oxphalin, among other phenolic azomethines, have been proven as strong inhibitors of lipoxygenases (LOX) from reticulocytes, soybean, thrombocytes as well as of quasi-LOX (hemoglobin) with IC50 values which correspond with those of known LOX inhibitors. The 5-LOX is likewise strongly, the cyclooxygenase of sheep seminal vesicles only weakly inhibited. Nevertheless, antiinflammatory effectivity was found in some carrageenin-induced inflammatory models of the rat as well as in the arachidonic acid- and croton oil-induced ear oedema of the mouse. Adjuvant arthritis, experimental pain, skin permeability and allergy models (anaphylactic paw oedema, cutaneous anaphylaxis, asthma, picryl chloride ear oedema) were not, only weakly or irregularly influenced. In the guinea pig ileum a certain antihistaminic, anticholinergic and leukotriene antagonistic activity was found. An inflammation-induced vasodepression (anaphylactic shock, dextran paw oedema. UV irradiation) was dose-dependently prevented or even reversed, obviously on the basis of oxygen radical scavenging.

[Inhibition of arachidonic acid-induced ear edema in the mouse with lipoxygenase-, cyclo-oxygenase- and dual inhibitors]. / Hemmung des Arachidonsäure-induzierten Ohrödems der Maus durch Lipoxygenase-,Cyclooxygenase- und Dualhemmer.

The ear edema of the mouse induced by local application of arachidonic acid is suitable for in vivo differentiation of lipoxygenase (LOX) and cyclooxygenase (COX) inhibitors. LOX blockers inhibit initially and plateau-like during the first hour of the course of the edema, while COX-blockers do so mostly only initially. For practice we recommend the measurement about 60 min following edema provocation. The dual blocker of LOX and COX, BW 755 c, acts like a LOX inhibitor.

Action of metal chelators on lipoxygenases, cyclooxygenase and on inflammation-induced vasodepression.

Metal complexing agents could exert antiinflammatory activity by inhibition of oxygen radical production, by inhibition of eicosanoid synthesis and by inhibition of metalloenzymes. We found a moderate antiedema activity of the iron (II)/iron (III)-chelator combination, o-phenanthroline/desferrioxamine, which could refer to an involvement of iron ions in the inflammatory reaction. The newly synthetized complexing agent ethylene-diimino-dibutyric acid caused weak antiedema and antivasodepressor effects which remain to be explained. Altogether, the in vivo effects of the metal chelators were rather weak. In vitro, only desferrioxamine produced a remarkable inhibition of two lipoxygenases.

Role of eicosanoids in inflammatory reactions of rats.

Nonsteroidal antiinflammatory agents of several subgroups, regarding eicosanoid-producing enzyme inhibition, have been tested in vitro and in vivo. There was no overt correlation between cyclooxygenase and/or lipoxygenase inhibition in vitro and antiinflammatory effects. It is possible that eicosanoids do not play a key role as inflammatory mediators, i.e. that the course of the life-protecting inflammatory reaction is ensured by an ensemble of mediators, cellular events and further mechanisms of the whole living organism.

Action of cyclooxygenase (COX) and lipoxygenase (LOX) inhibitors as well as of oxygen free radical scavengers (OFRS) in the inflammation-induced vasodepression.

Vasoprotective activity of COX inhibitors (diclofenac, piroxicam, indomethacin, aspirin) and of LOX inhibitors (nordihydroguaiaretic acid, propyl gallate, oxphaman) or of both (BW 755c) as well as of OFRS (mannitol, ethanol, thiourea, vitamin E) was investigated ex vivo in the isolated perfused hindlegs of the animals after in vivo u.v.-irradiation and antigen-provocation in mice and rats, respectively. The results show that vasodepression is mainly prevented by OFRS, antioxidants/LOX inhibitors and aspirin, but not by other COX inhibitors.

Carrageenin-induced thrombosis in rats and mice: a model for testing antithrombotic substances?

Among different carrageenins, kappa-carrageenans were found to be thrombogenic, whereas lambda-carrageenins were inactive in this respect, although the latter substances exerted a stronger edemogenic activity. Kappa-carrageenin (Sigma) was the most potent thrombogen. As the consequence of thrombosis tail infarction became visible some minutes after i.v. administration, but it was delayed for about 3 hours after the i.p. route and for about 6 hours after subplantar injection. Infarction frequency as well as extent of infarction was inhibited by heparin, cyproheptadine, phentolamine and dibenamine. Other substances like aspirin and dipyridamole showed no or only weak effects. Advantages of the carrageenin-induced thrombosis model in rats and mice are: (i) simple induction in small laboratory animals, (ii) easy observation and quantification all the time without killing the animals, and (iii) possible external testing of antithrombotic agents by applying substances on the tail.

Carrageenin-induced thrombosis in the rat and mouse as a test model of substances influencing thrombosis.

Kappa-carrageenins cause disseminated intravascular coagulation with thrombosis of the tail in rats and mice. Frequency and extent of tail thrombosis were used for determining antithrombotic effects after systemic and local external administration of substances. Inhibitors of cyclooxygenase and thromboxane synthetase caused irregular inhibition of thrombosis after relatively high doses. The cyclooxygenase/lipoxygenase inhibitor BW755C was ineffective after 50 mg/kg. Hitherto, no effective substances could be found after external administration on the tail of mice. At present, no convincing explanation for thrombogenic activity of kappa-carrageenin can be given. The advantages of the thrombosis model are discussed.

Blood vessel reactivity on noradrenaline, vasopressin, and prostaglandin F2 alpha, resp., in the isolated perfused hind legs of rats with edemas or adjuvant arthritis.

In the isolated perfused hind legs of rats with enemas induced by carrageenin, dextran or Freund's adjuvant in both paws, resting perfusion pressure was slightly increased whereas the vasopressor action of noradrenaline, lysine-vasopressin and prostaglandin F2 alpha, was decreased. Admixture of indomethacin (3 micrograms.ml-1) to the perfusion fluid led to a decrease of resting perfusion pressure whereas its influence on EAmax of noradrenaline was only weak under these conditions. Concomitantly, the contents of prostaglandin E-like substances in the perfusate decreased. Prostaglandin F2 alpha exhibits only weak vasopressor activity in isolated perfused hind legs of rats with carrageenin edema. Altogether, the effect of indomethacin on resting perfusion pressure as well as on EA and EAmax, resp., of agonists is difficulty to explain by its effect on arachidonic acid cascade. The pD2-value of noradrenaline (4.68 - 4.87) and lysine-vasopressin (6.02 - 6.04) was apparently not changed, at least not decreased, in the acute phase of inflammation indicating no impairment of receptor affinity of noradrenaline and vasopressin in inflammation. Blood vessel reaction is apparently influenced in inflammation mechanically by the increased tissue pressure as well as by molecular mechanisms consisting in the presence of inflammatory mediators and/or particularly in a reduced intrinsic sensitivity of the blood vessel muscle itself.

Effect of lysolecithin on blood vessel reactivity and of some ethers and esters of glycerophosphocholine and phosphocholine, respectively, on aggregation of rat platelets.

Infusion of lysolecithin (LPC; e.g. 88 microgram/ml for 0.5-1.0 min) did not significantly impair the vasopressor action of norepinephrine (NE), prostaglandin F2 alpha (PGF2 alpha) and extract of posterior pituitary (EPP) in the isolated perfused hind legs of rats. In other words, vascular smooth muscle behaves differently from the smooth muscle of the guinea-pig small intestine, since, in the latter, contractions evoked by acetylcholine, prostaglandins etc., are inhibited by LPC. Triton X 100 which, by comparison, was used as a detergent effective on the guinea-pig small intestine, depressed the vasopressor effect of NE, PGF2 alpha and EPP. LPC, at low concentrations (40 mumol/l), potentiated (15% max.) ADP-induced platelet aggregation (PA) in rat PRP but, at high concentrations, inhibited PA (IC50 = 390 mumol/l). 2-Hexadecylglycerophosphocholine and its short-chain 1-alkyl ethers, which are structurally related to platelet-activating factor, as well as some long-chain alkanol phosphocholine esters, were somewhat more active than LPC. Dipalmitoyllecithin (4-700 mumol/l) was without any effect.

Prostaglandin release from perfused, isolated hind legs in normal and arteriosclerotic rats.

Intraarterial injection of 10 microgram noradrenaline (NA) produced a stronger increase of prostaglandin E-like material (PGE) in the outflow of isolated perfused hind legs of normal rats than of those of arteriosclerotic rats. The vasopressor effect of NA in normal rats was stronger than in arteriosclerotic rats, which could be explained on the basis of PGE release.

[Peroxidase activity in the paw of animals in adjuvant arthritis and its changes by antiphlogistics in vitro]. / Peroxidaseaktivität in der Pfote bei Adjuvansarthritis und ihre Beeinflussung durch Antiphlogistika in vitro

The peroxidase activity is increased in the inflamed paw of rats with adjuvant arthritis. The increase is biphasic like for other enzymes. The higher peak occurs during primary inflammation in the injected paw according to the behaviour of polymorphonuclear leucocytes. Of 32 substances tested in vitro, among them 19 antiphlogistics and derivatives, especially the antiphlogistics inhibited the peroxidase reaction. Therfore it seems not unlikely that inhibition of the peroxidase reaction is involved in the mechanism of activity of anti-inflammatory agents.

[Modification of the adjuvans arthritis by carrageenin, compound 48/80, histamine- and serotonin antagonists, non-steroid antiphlogistics as well as protease inhibitors and their possible relations to inflammation mediators]. / Beeinflussung der Adjuvansarthritis durch Carrageenin, Compound 48/80, Histamin- und Serotoninantagonisten, nichtsteroidale Antiphlogistika sowie Proteasenhemmstoffe und deren mögliche Beziehungen zu Entzündungsmediatoren

1. Injections of carrageenin (1,25 mg/kg i.v.) from the 1st to the 3rd day and then each 2nd or 3rd day inhibited paw swelling in adjuvant arthritis of the rat during the time of treatment. Injections from the 11th to the 15th day were ineffective. The level of plasma kininogen was slightly decreased but the total complement serum level was significantly lowered. 2,5 and 3 mg carrageenin/kg respectively were toxic after repeated injections. After a single administration the levels of plasma kininogen and of total serum complement were decreased by 50% although paw swelling was not affected. 2. Pentosane polysulfoester (25 mg/kg i.v.) did not influence paw swelling despite daily administration from the 1st to the 17th day. Heparin (10 000 IE/kg i.v.) was likewise ineffective. 3. Single or repeated injections of compound 48/80 (0,125-0,5 mg/kg i.v.; 1-5 mg/kg i.p.; 3-6 mg/kg s.c.), reserpine (0,2 mg/kg i.p.), cyproheptadine (5 mg/kg i.v.), bromolysergic acid diethylamide (2 x 2 mg/kg i.v.) or metiamide (10 mg/kg i.v.) were without effect on paw swelling. Neither did compound 48/80 effect the complement serum level. 4. Daily administration of chloropromazine (4-10 mg/kg p.o.) or of promethazine (10-15 mg/kg s.c. or p.o.) inhibited paw swelling in the first phase of adjuvant arthritis but not in the second one. 5. The soybean trypsin inhibitor (15 mg/kg i.v.) inhibited paw swelling significantly up to the 4th day, the Kunitz inhibitor (25 000 E/kg i.v.) was ineffective. 6. The content of prostaglandin E of the inflamed paws was increased threefold in both phases of arthritis. The results are discussed with regard to the putative role of mediators of inflammation (histamine, serotonin, kinins, prostaglandins, lysosomal enzymes, lymphokines, complement).