J. R. Walker

Anti-inflammatory drugs, prostaglandins and leucocyte migration

The action of some non-steroidal acidic anti-inflammatory drugs, aspirin, phenylbutazone and indomethacin, in reducing leucocyte migration into the exudates of inert porous sponges implanted subdermally in the rat has been shown to be distinct from their effect in reducing the content of prostaglandins in the exudates. It is concluded that a component of the anti-inflammatory and antirheumatic actions of the drugs is concerned with a mechanism other than inhibition of prostaglandin biosynthesis.

THE EFFECTS OF SOME ANTIRHEUMATIC DRUGS ON AN in vitro MODEL OF HUMAN POLYMORPHONUCLEAR LEUCOCYTE CHEMOKINESIS

1 A rapid, reproducible in vitro assay for studying the chemokinetic movement of human polymorphonuclear leucocytes (PMNs) is described. Two synthetic peptides, formyl methionyl‐leucylphenylalanine (FMLP) and formyl methionyl‐phenylalanine (FMP), were used as standard chemokinesins. 2 Maximal chemokinetic movement was observed with peptide concentrations of 2.5 nm (FMLP) and 100 μm (FMP). EC50 values of 650.0 ± 60.0 pm and 27.0 ± 3.5 μm respectively are similar to those reported for chemotactic activity of the peptides in micropore filter assays. 3 The PMN chemokinetic response to FMLP was enhanced by histamine (100 nm) and vitamin C (2.5 μm). 4 Human serum albumin was shown to induce chemokinesis but to antagonize the response to FMLP in a dose‐related fashion. Fibrinogen similarly antagonized the cell response to peptide. 5 Levamisole (250 nm to 2.5 μm) significantly potentiated the chemokinetic responses to FMLP and FMP in a dose‐related manner. The chemokinetic response to FMLP was unaffected by d‐penicillamine (250 μm to 10 mm) while alclofenac (500 μm to 1 mm), salicylic acid (250 μm to 10 mm) and indomethacin (100 μm to 1 mm) caused dose‐related inhibition.

PGI2: a potential mediator of inflammation.

PGI2, but not its metabolite 6oxoPGF1alpha, is equivalent in potency to PGE1 as a potentiator of carrageenan, histamine and bradykinin-induced rat paw oedemas. PGI2 must, therefore, be considered as a potential mediator of inflammatory processes.

Separate anti-inflammatory effects of indomethacin, flurbiprofen and benoxaprofen

An important query about non-steroidal anti-inflammatory drugs is whether they have either a common single mode of action or different multiple interactions with various aspects of inflammatory responses. An inhibitory action on prostaglandin biosynthesis has been advanced as a mechanism for the first possibility (Vane, 1973). More recently evidence has accumulated that the drugs possess anti-inflammatory effects which appear to be independent of prostaglandin systems (Bonta, Bult & others, 1977; Crook, Collins &others, 1976). One of the most relevant is an interference with the emigration of leucocytes into inflammatory sites (Walker, Smith & Ford-Hutchinson, 1976a). In the present work the effects of three acidic non-steroidal drugs, indomethacin, flurbiprofen and benoxaprofen, on the production of prostaglandins and the migration of leucocytes into an inflammatory exudate in vivo, have been studied. All three drugs possess a similar spectrum of experimental anti-inflammatory activity (Glenn, Rohloff & others, 1973 ; Adams, McCullough & Nicholson, 1975; Cashin, Dawson & Kitchen, 1977) but differ in that the first two are potent inhibitors of prostaglandin synthetase activity in vitro (Crook & Collins, 1975) whereas benoxaprofen is only a weak inhibitor of the enzyme system (Cashin & others, 1977). Indomethacin was obtained from Merck Sharp and Dohme. Benoxaprofen (2-[4-chlorophenyl]-cc-methyl-5benzoxazone acetic acid) was obtained from Dr W. Dawson, Lilly Research Centre, Ltd, Windlesham, Surrey, England, and flurbiprofen (2-[fluoro-4-biphenylyl] propionic acid) was obtained from Dr S. Adams, Boots Drug Co. Ltd, Nottingham, England. The 9 h sponge implantation technique, the estimation of leucocyte migration and prostaglandin-like activity in the sponges were as described previously (Walker & others, 1976a). Drugs were administered orally as a suspension in Tween 80 to groups, each of 5 rats, 1 h before sponge implantation. The effects of the three drugs on prostaglandin accumulation and leucocyte migration into the 9 h sponge exudates are shown in Table 1 . Indometha& and flurbiprofen, which are potent inhibitors of prosta. glandin synthetase in vitro, strongly inhibited the accumulation of prostaglandin activity in vivo in the sponges. This inhibition occurred at lower dosages than those required to inhibit white cell migration in the system and well below those required to inhibit other experimental models of inflammation (Glenn & others, 1973; Adams & others, 1975). Benoxaprofen only showed inhibition of prostaglandin production in vivo at doses equivalent to those required to inhibit white cell migration in the model and similar to those needed to affect other experimental models (Cashin & others, 1977). These findings demonstrate a lack of correlation

Effects of a human plasma fraction on leucocyte migration into inflammatory exudates.

A fraction prepared from normal human plasma inhibits the migration of polymorphonuclear and mononuclear leucocytes into inflammatory exudates produced by the intrapleural injection of carrageenan or turpentine and by the subcutaneous implantation of polyvinyl sponges in the rat. The mechanism of the effect does not involve complement depletion.

Assessment of anti-inflammatory activity by sponge implantation techniques

Abstract A technique for implanting inert porous sponges subdermally in the rat is described. The model of acute inflammation is particularly useful because it allows the easy collection and examination of both cellular and fluid phases of the exudate formed within the sponge. It may be modified to study more chronic inflammatory responses, including the formation of granulomata, and also as a model of delayed hypersensitivity thus allowing the anti-inflammatory activity of several types of antirheumatic drugs to be investigated.

Prostaglandins and leucotaxis

Freshly prepared solutions of prostaglandin E1 exhibit chemotactic activity against rabbit polymorphonuclear leucocytes harvested from the peritoneal cavity but are devoid of such activity either against similar cells obtained from the rat or against polymorphonuclear leucocytes obtained from the blood of the rabbit, rat and man. Some implications of these findings with respect to the development of inflammatory responses and the mode of action of non‐steroidal acidic anti‐inflammatory drugs are discussed.

Platelets, prostaglandins and inflammation

In exudates of implanted sponges in rats, made thrombocytopenic by the administration of anti-platelet serum, there are significant reductions in the platelet and leucocyte counts and of the content of prostaglandin-like activity. It is concluded that platelets migrate into the developing sponge exudates, are the source of the prostaglandins and interact with complement to cause leucocyte migration. In normal animals the administration of indomethacin and sodium salicylate cause similar effects to thrombocytopenia whereas the injection of human plasma fraction affects only the leucocyte migration. One of the sites of the anti-inflammatory action of conventional non-steroidal drugs may be concerned with the migration of platelets into inflammatory lesions.

THE EFFECTS OF d‐PENICILLAMINE AND LEVAMISOLE ON LEUCOCYTE CHEMOTAXIS IN THE RAT

1 The administration of D‐penicillamine (25 mg/kg) or levamisole (5 mg/kg) had no effect on leucocyte emigration into the exudates formed in inert sponges implanted in normal rats. 2 In rats, previously sensitized to Bordetella pertussis and implanted with sponges containing pertussis vaccine, an increased leucocyte migration into the exudates occurred; this was significantly enhanced by the administration of the drugs. 3 Neither drug in vitro affected the chemotaxis of rat polymorphonuclear leucocytes although random migration was significantly increased by levamisole (2 μg to 1 mg/ml). Neither drug affected the chemotaxis of rat mononuclear cells although levamisole (25 μg/ml) significantly increased that of human monocytes. 4 It is concluded that both drugs produce similar effects in an animal model of delayed hypersensitivity and that their clinical antirheumatic actions may have common elements.

Aspirin, salicylate and prostaglandins

The effects of aspirin, salicylic acid and gentisic acid on the paw swellings in the arachidonic acid-potentiated and in the conventional carrageenan-induced oedema tests as well as on the content of prostaglandin-like activity and leucocyte migration in the exudate of inert implanted sponges in the rat have been studied. It is concluded that aspirin exerts two separate inhibitory effects on prostaglandin formation in vivo, a rapid action of the intact molecule on easily accessible tissues and a later action due to its metabolic conversion to salicylic acid. Salicylic acid inhibits prostaglandin biosynthesis in vivo as the salicylate ion itself and there is no formation of a subsequent ‘active’ metabolite.