Formation Of Slow Reacting Substance Research Articles

Immune complex mediated inflammation in the mouse peritoneal cavity: A method for investigating inhibitors of arachidonate metabolism

Intraperitoneal injection of washed, preformed ovalbumin/anti-ovalbumin immune complexes caused the production of slow-reacting substance (SRS), prostaglandin E 2, 6 keto prostaglandin F 1a, vascular permeability (measured as extravazation of pontamine sky blue injected previously into the bloodstream), and polymorphonuclear leukocyte (PMN) infiltration in the mouse peritoneal cavity. Arachidonic acid metabolites appeared early in the response at 7.5 min; dye extravazation and PMN infiltration peaked at 30 min and 4 hr, respectively, after immune complex injection. BW755C and phenidone given orally 30 min before immune complex injection inhibited dye extravazation and SRS formation at 7.5 and 30 min. Indomethacin by the same regimen inhibited dye extravazation at 7.5 min but not at 30 min, while enhancing SRS formation at both times. Dexamethasone inhibited dye extravazation and to some extent SRS formation at both times. The data suggests that the mouse model may be useful for demonstrating an antiinflammatory activity of dual inhibitors that is clearly distinguishable from the activities shown by classical nonsteroidal antiinflammatory drugs and steroids.

Relationship of biosynthesis of slow reacting substance to intracellular glutathione concentrations.

To further elucidate the role of glutathione (GSH) in the biosynthesis of slow reacting substance (SRS), SRS generation was studied in rat basophilic leukemia cells that had been preincubated with 2-cyclohexen-1-one or diethyl maleate to decrease their intracellular GSH concentrations. At low GSH levels SRS formation was markedly inhibited. The formation of other lipoxygenase products was much less affected, although some decrease in 5-hydroxyicosatetraenoic acid formation also occurred, apparently due in part to less rapid reduction of the 5-hydroperoxide.

Effect of the 5-hydroperoxide of eicosatetraenoic acid and inhibitors of the lipoxygenase pathway on the formation of slow reacting substance by rat basophilic leukemia cells; direct evidence that slow reacting substance is a product of the lipoxygenase pathway.

Previous studies in a line of rat basophilic leukemia (RBL 1) cells have indicated that the slow reacting substance (SRS) made during stimulation with the divalent cation ionophore, A23187, is derived from arachidonic acid (AA). In the present report, various inhibitors of AA metabolism were compared with regard to their effects on SRS formation and incorporation of radioactivity from [1-14C]-AA into known metabolites of the lipoxygenase and cyclooxygenase pathways. An apparently close parallel between lipoxygenase product formation and SRS synthesis is demonstrated. In addition, exogenous 5-hydroperoxy-eicosatetraenoic acid (5-HPETE) has been shown to markedly enhance SRS synthesis, even when A23187 is absent. The data provide very strong evidence that SRS is produced through the lipoxygenase pathway.

Precursor role of arachidonic acid in release of slow reacting substance from rat basophilic leukemia cells.

The release of slow reacting substance (SRS) from rat basophilic leukemia cells (RBL-1) by the ionophore A23187 (5-10 mug/ml) was stimulated 5-fold by arachidonate and inhibited 78% by 5,8,11,14-eicosatetraynoate (an inhibitor of both fatty acid cyclooxygenase and lipoxygenase). Linoleic acid and linolenic acid both inhibited SRS formation, whereas indomethacin (a cyclooxygenase inhibitor) had no effect. Radiolabel from [14C]- or [3H]arachidonate was incorporated into SRS as indicated by comigration of radioactivity and bioreactivity in several chromatographic systems after purification to apparent radiochemical homogeneity. The radiolabeled SRS was clearly separated chromatographically from other known arachidonate metabolites. Thus, SRS appears to be a previously undescribed product of arachidonic acid metabolism, probably formed through the lipoxygenase pathway. The ability to prepare purified, biosynthetically labeled, SRS should be of considerable help in further studies of its structure, biologic function, and catabolism.

Ca++ dependence of histamine release and formation of slow reacting substance in the cat paw.

AbstractThe influence of divalent cations on the release of histamine and the formation of slow reacting substance (SRS) in the perfused cat paw induced by compound 48/80 was studied. Ca++ was found to have a stimulatory effect on both processes. Ca++ had no releasing effects per se. When the paws were perfused for 1 or 2 hrs with Ca++‐free salt solution, prior to compound 48/80, the release was depressed. Histamine release and formation of SRS were inhibited by disodium ethylene diamine tetraacetate (EDTA). In both instances the EDTA‐blockade could be reversed by Ca++, whereas Ca++, Mg++, Mn++ and Ni++ were ineffective substitutes. The EDTA‐blockade of histamine release but not that of the formation of SRS was also reversed by Sr++ and in some experiments by Ba++. The stimulatory effect of Sr++ on the release of histamine was weaker than that of Ca++. No significant stimulatory effect of Sr++ on the formation of SRS was observed. It is concluded that the release of histamine and the formation of SRS in the cat paw are Ca++‐dependent processes although in the case of histamine release Ca++ can partially be replaced by Sr++.

Release of histamine and formation of slow reacting substance in the cat paw induced by compound 48-80.

AbstractA method for perfusion of cat paws with constant flow and temperature is described. This technique has been used to study the relationship between the efflux of histamine and slow reacting substance (SRS) from cat paws perfused with compound 48/80. The threshold dose of compound 48/80 was 0.001–0.01 μg/ml for both histamine and SRS releasing effects. No correlation between the yields of histamine and SRS was noted. On repeated administration of compound 48/80 the histamine releasing effect declined and the output of SRS was cor respondingly reduced. The release of histamine and the formation of SRS were inhibited by ninhydrin, N‐ethylmaleimide and potassium cyanide indicating that both processes are dependent on energy‐requiring enzymatic reactions. Apart from their antagonism of the histamine releasing effect of compound 48/80, ninhydrin and N‐ethylmaleimide released histamine per se. No SRS was detected in the effluents from paws perfused with these agents alone. The yields of both histamine and SRS were higher at 22o C than at 37oC. Regardless of temperature the efflux of histamine consistently preceded that of SRS. A temperature dependent inactivation of SRS by cat serum was demonstrated. The results suggest that the initial reaction(s) activated by compound 48/80 is common for the release of histamine and the formation of SRS in the cat paw but that these processes then proceed independently.

Effect of nicotinamide on in vitro histamine release and slow-reacting substance formation in anaphylactically sensitized lung and its relation to acid-soluble nucleotides

The present experiments confirm that nicotinamide partially inhibits the in vitro release of histamine from anaphylactically sensitized guinea pig lung on incubation with antigen. It has also been demonstrated that formation of SRS-A is reduced by nicotinamide. No change in DPNase activity of lung tissue was demonstrable after in vitro anaphylaxis. In vitro anaphylaxis caused a 28 to 47 per cent reduction in lung content of acid-soluble nucleotides as measured by the orcinol reaction. The possible significance of this change is discussed.