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Injections of heparin had no effect on the dextran anaphylactoid reaction in rats but neutralised the inhibitory action both of protamine and of borate. Heparin, protamine, and borate did not modify the histamine or 5-hydroxytryptamine responses in rat paws and so their effects on dextran may be due to modification of the release mechanisms of these two amines from tissue mast cells. Correspondence to: Dr. G.B. West, Department of Paramedical Sciences, North East London Polytechnic, Romford Road, London E15 4LZ (England) Almost 20 years ago, heparin was reported [2] to increase the rate of wound healing in rat skin, with even better results when histamine was administered just before the heparin. Protamine, on the other hand, had no effect on wound healing by itself but it neutralised the increase in tensile strength of skin wounds produced both by heparin and by heparin and histamine. Protamine is considered to be a selective antagonist of heparin in many situations. Heparin also stimulates the formation of cartilage and antagonises some of the actions of glucocorticoids. It was of interest, therefore, to test heparin and protamine in an acute inflammatory reaction such as the dextran anaphylactoid oedema in rats, in which histamine and 5-hydroxytryptamine are first released from mast cells and heparin release follows. Recently, we have shown [4] that sodium salicyiate enhances the dextran anaphylactoid response in rats administered with the dextran. This followed the report [3] that salicylates are effective adjuvants for the rectal absorption of insulin (a polypeptide) and heparin (a mucopolysaccharide), compounds which normally are either degraded or poorly absorbed after oral administration. Sodium borate has also been shown [1] to enhance histamine release from isolated rat peritoneal mast cells induced by palytoxin, a long aliphatic chain with interspersed cyclic ether, hydrox-yl and carboxyl groups, isolated from marine coelen-terates. Salicyiate and borate were therefore also tested on the dextran anaphylactoid reaction in rats in the presence and absence of heparin. In an attempt to identify their mode of action, the effects of heparin, protamine, borate and salicyiate on local actions of histamine and 5-hydroxytryptamine in rat paws were also studied. Groups of 5 Wistar rats (200–300 g) were obtained from the Tuck colony and injected with clinical dextran (Intradex, molecular weight 110,000) to produce the anaphylactoid reaction after intraperitoneal injection (200 mg·kg-1) or intrapedial injection (100 μg). The responses were measured as increases in hind paw volume as determined on a volume differential meter over 5 h. 7 days later, heparin, protamine, sodium borate or sodium salicyiate were included with the dextran or injected 30 min before or after it. The results shown in the figures are the means ± SEM, and statistical analysis of the results was by the Student’s t test. The simultaneous administration of doses of heparin up to 2,000 IU· kg-1 by the intraperitoneal route did not modify the increase in hind paw volume produced by dextran. However, protamine D ow nl oa de d by : 54 .7 0. 40 .1 1 10 /5 /2 01 7 9: 41 :3 5 P M significantly reduced the reaction in a dose-dependent fashion, as shown in figure 1. When heparin was included in the dextran-protamine mixture, the inhibitory action of protamine was prevented. This result was expected as protamine is usually a selective antagonist of heparin. When sodium borate was tested in a similar manner, it too significantly reduced the dextran anaphylactoid reaction when used in doses of 0.5–2 mg·kg-1 and again heparin significantly antagonised the inhibitory action (fig. 2). This result was unexpected as 374 West Time after dextran, miπ Fig. 1. Effect of protamine (O, 0.5 mg·kg”‘; •‚ 2 mg·kg-1) on the increase in hind paw volume of rats produced by clinical dextran (200 mg·kg”‘. All doses injected intraperitoneally as mixtures. Results shown as mean percent increases ( ± SEM) in groups of 5 rats. Control responses without protamine (Δ) and the effect of he-parin (A, 2,000 IU·kg”1) on the protamine inhibition (2 mg·kg-1) are also shown. Note the dose-dependent inhibition of the dextran response by protamine and its antagonism by heparin. D ow nl oa de d by : 54 .7 0. 40 .1 1 10 /5 /2 01 7 9: 41 :3 5 P M Time after dextran, minFig. 2. Effect of sodium borate (·, 2 mg·kg”1) on the increase in hind paw volume of ratsproduced by clinical dextran (200 mg· kg”1). All doses injected intraperitoneally as mixtures.Results shown as mean percent increases ( ± SEM). Control responses without borate (Δ) and theeffect of heparin (A, 2,000 IU·kg-1) on the borate inhibition are also shown. Again, note theinhibition of the dextran response by borate and its antagonism by heparin.Fig. 3. Effect of protamine (·, 4 μg) on the increase in hind paw volume of rats produced byclinical dextran (100 μg). All doses injected intrapedially as mixtures. Results shown as meanpercent increases ( ± SEM) of groups of 5 rats. Control responses without protamine (Δ) and theeffect of heparin (A, 20 IU) on the protamine inhibition are also shown. Note the inhibition byprotamine of the dextran response and its antagonism by heparin.borate enhances the action in vitro of palytoxin, one of the most potent animal toxins known, butdoes not enhance histamine release by compound 48/80 or concanavalin A [1]. When injected 30min after (but not 30 min before) the dextran, both protamine and borate exerted their inhibitoryeffects. Sodium salicy-late, as expected, enhanced the dextran anaphylac-toid oedema in ratswhen used in doses of 30 mg·kg-1, but this enhancement was not modified by heparin.When the drugs were administered locally into rat paws, protamine (4 μg) and borate (4 μg)reduced the dextran response (100 μg), and heparin (20 IU) antagonised these effects. The resultusing protamine is shown in figure 3. Saliycylate enhanced the local dextran reaction but heparindid not change this enhancement. The local oedema reactions produced in rat paws by histamine(50 μg) and 5-hydroxytrypta-mine (1 μg) were not modified by heparin, protamine, borate orsalicylate. It is possible, therefore, that most of these effects on dextran are due to a modificationof the release mechanisms of histamine and 5-hy-droxytryptamine from tissue mast cells. Theimportance of the plasma membranes of these cells is again stressed.Heparin and the Dextran Anaphylactoid Reaction in Rats375 ReferencesChhatwal, G.S.; Ahnert-Hilger, G.; Beress, L.; Habermann, E.: Palytoxin both induces andinhibits the release of histamine from rat mast cells. Int. Archs Allergy appl. Immun. 68: 97–100(1982).Fenton, H.; West, G.B.: Studies on wound healing. Br. J. Pharmacol. 20: 507–515(1963). Downloadedby: 54.70.40.11-10/5/20179:41:35PM Nishihata, T.; Rytting, J.H.; Higuchi, T.; Caldwell, L.: Enhanced rectal absorption of insulin andheparin in rats in the presence of non-surfactants. J. Pharm. Pharmac. 33: 334–335 (1981).4 West, G.B.: Enhancement of the anaphylactoid reactions in rats by salicylates. Int. ArchsAllergy appl. Immun. 68: 185–187 (1982). Downloadedby: 54.70.40.11-10/5/20179:41:35PM