Inhibition of IgE-mediated allergic histamine release from rat peritoneal mast cells by azelastine and selected antiallergic drugs

The ability of azelastine to inhibit IgE-mediated allergic histamine release from the peritoneal mast cells of actively sensitized rats was investigated and compared with selected antiallergic agents. Azelastine added simultaneously with the allergic stimuli (ovalbumin, OA, 10 g/ml + phosphatidylserine, PS, 10 g/ml) or preincubated with cells for 10 min prior to antigen challenge produced similar concentration-dependent inhibition of allergic histamine release. The IC_50s (M) following 10-min preincubation were as follows: azelastine = 4.8; astemizole = 86.3; ketotifen = 112.2; diphenhydramine = 133 and theophylline = 2040.3. At IC₅₀ level azelastine was about 18, 23, 28 and 425 times as effective as astemizole, ketotifen (newer histamine H₁-receptor antagonists), diphenhydramine (a traditional H₁-receptor antagonist), and theophylline (a phosphodiesterase inhibitor), respectively. Sodium cromoglycate in a concentration range or 1–1000 M (0 or 10-min preincubation) failed to exert any inhibitory effect. These data showed that among six drugs tested azelastine is the most potent inhibitor of allergic histamine release from rat peritoneal mast cells.     

Inhibition of allergic histamine release from rat peritoneal mast cells by azelastine. Interaction with selected antiasthmatic drugs

The in vitro interaction of azelastine (a new antiallergic/antiasthmatic drug) with albuterol (a beta 2 bronchodilator), theophylline (a phosphodiesterase inhibitor), disodium cromoglycate (DSCG, a mast cell-stabilizing agent) and prednisolone (a steroid) was studied for effects on allergic histamine release from rat peritoneal mast cells (RPMCs). The RPMCs preincubated with albuterol, theophylline, DSGC (10 min) and prednisolone (2h) caused a 2- to 18-fold decrease in the IC30 for azelastine. Significant potentiation (synergism) was seen only with albuterol (0.01, 0.1 and 1.0 microM, 10 min) and theophylline (1.0 microM, 10 min). The pre-exposure of RPMCs with DSCG (0.1, 1.0 and 10 microM) for a period of 10 min produced a slight leftward shift of azelastine's concentration-effect curve (0.01, 0.1 and 10 microM added immediately before antigen challenge). This effect was not dependent on the concentration of DSCG. These data demonstrated (1) the lack of cross-tachyphylaxis between DSCG and azelastine, and (2) the synergistic interaction between azelastine and albuterol or theophylline.     

The inhibition of histamine release by antiallergic drugs.

Three new antiallergic drugs, Doxantrazole, PRD-92 and N5', as well as disodium cromoglycate, inhibited the IgE-mediated PCA reaction in the rat triggered by the homologous antigen, but did not have an antagonistic effect on histamine itself. Moreover, all the drugs examined caused in vitro inhibition of antigen-mediated histamine release from peritoneal mast cells and chopped lung tissue of sensitized rats producing IgE antibodies. Doxantrazole had a synergistic effect on the inhibition of histamine release by isoproterenol, whereas the other drugs were devoid of this capacity. PRD-92 and N5' inhibited the ionophore A23,187 induced histamine release, but did not have any effect on the D2O-enhanced histamine release which was triggered by antigen.     

Inhibition by azelastine of nonallergic histamine release from rat peritoneal mast cells

The ability of azelastine and selected antiallergic drugs to inhibit compound 48/80-induced and PS-potentiated, Con A-induced histamine release from RPMC was investigated. Azelastine, ketotifen, theophylline, and DSCG added simultaneously with the secretagogues or preincubated with the RPMC for 10 min before the addition of secretagogues produced concentration-dependent inhibition of histamine release. In general, the relative order of potency at calculated IC50 level was as follows: azelastine greater than ketotifen greater than theophylline greater than DSCG. The preincubation of RPMC with azelastine for 10 min exerted 3.5 times greater inhibition of Con A plus PS-stimulated histamine release but did not influence the inhibitory activity on compound 48/80-induced release. The duration of preincubation did not influence the inhibitory effects of ketotifen with either secretagogue. Theophylline and DSCG exerted significantly greater inhibition when they were added simultaneously with Con A plus PS. The inhibitory activity of DSCG was also significantly improved upon simultaneous addition with compound 48/80. These data demonstrated that azelastine is the most potent inhibitor of nonallergic histamine release from RPMC among the four antiallergic drugs examined.     

Augmentation of allergic histamine release from human leukocytes by nonsteroidal anti-inflammatory-analgesic agents

Augmentation of allergic histamine release in vitro with human leukocytes was produced by numerous nonsteroidal anti-inflammatory--analgesic agents, primarily the arylalkanoic and anthranilic acids. Augmentation occurred without release of histamine by the agents in the absence of the allergen (ragweed) and only under conditions of an accompanying release by the allergen. As a consequence of augmentation, less allergen was necessary to produce a given response in the presence of these agents. It is suggested that some of these agents might enhance mediator release in immediate-type hypersensitivity reactions. The same agents reported to exacerbate chronic urticaria, i.e., indomethacin, mefenamic acid, sodium salicylate, aspirin, sodium benzoate, and tartrazine, also augmented allergic histamine release. Pharmacologically mediated augmentation of mediator release from stimulated cells is suggested to be involved in the exacerbation of existing chronic urticaria by the acidic nonsteroidal anti-inflammatory-analgesic agents.     

Intracellular calcium release induced by histamine releasers and its inhibition by some antiallergic drugs

When rat mast cells loaded with fluorescent Ca2+ indicator Quin 2 were exposed to either compound 48/80 (0.1 micrograms/mL) or substance P (2 microM) at 37 degrees C for 30 seconds in a Ca-free medium, a marked increase of Quin 2 fluorescence was noticed, indicating that Ca2+ was released from the intracellular Ca store. The pixel values of the whole cell image were displayed in a three dimensional projection. When mast cells were exposed to 48/80, the fluorescent increase was reflected as an increase of height and spreading of the image. When 0.01 to 1 mM of db-cAMP was pretreated for five minutes, an increase of Quin 2 fluorescence was inhibited in a dose-dependent fashion. Theophylline pretreatment also showed a preventive effect at 1 to 5 mM. A marked inhibition of the Quin 2 signal was induced by pretreatment with 0.01 mM of terfenadine (63.4% inhibition) or ketotifen (26.6% inhibition). Disodium cromoglycate also showed a similar inhibitory effect. In the measurement of the order parameters of liposomes, the addition of either terfenadine or ketotifen into the lipids increased the parameter value, indicating they provide the membrane stabilizing action.     

Nonsteroidal antiinflammatory drugs induce UV-dependent histamine and leukotriene release from peripheral human leukocytes

The effect of UV-A radiation on the in vitro release of vasoactive mediators from human peripheral leukocytes incubated with different nonsteroidal antiinflammatory drugs (NSAIDs) was studied in the newly developed photo basophil histamine release test (PBHRT). Washed peripheral leukocytes were incubated with 10(-6) to 10(-3) M of various NSAIDs and exposed to 1-100 J/cm2 UV-A. Maximum histamine release was 4% with acetylsalicylic acid, 10% with benoxaprofen, 20% with thiophene, 28% with diclofenac, 39% with tiaprofenic acid, 40% with carprofen, 55% with ketoprofen and not demonstrable with indoprofen. Maximal reactions occurred at UV-A doses of 25 or 50 J/cm2. Leukotriene B4 (LTB4) generation in the supernatants of cells treated with UV-A or NSAID alone was below or close to the detection limit (145 pg/ml). On the contrary, UV-A irradiation of cells incubated with NSAID led to marked LTB4 generation (up to 1,000 pg/ml). The results indicate that many NSAIDs can induce photosensitization in vitro, the PBHRT being a promising new method for identification of possibly phototoxic compounds. Release of vasoactive mediators like histamine or leukotrienes may be involved in the in vivo development of phototoxic or photoallergic side reactions.     

Inhibition of IgE-mediated histamine release from human leukocytes by a new xanthine derivative,D 4026

Pretreatment of human leukocytes with the new xanthine compound, 3,7-dihydro-1,8-dimethyl-3-phenyl-1H-purine-2,6-dione (D 4026), induced a dose-dependent and statistically significant inhibition of immunoglobulin E-mediated histamine release in the concentration interval 0·1–1000 M. Histamine release elicited with suboptimum amounts of the triggering agent (anti-IgE or antigen) was inhibited to a greater extent than a release initiated with optimum amounts. At a concentration of 10 M, D 4026 had at least the same inhibitory effect as 100 M theophylline. When leukocytes were incubated simultaneously with D 4026 and a histamine H-2 receptorstimulating drug (histamine or clonidine), the two drugs combined induced an inhibition significantly greater than the sum of their individual inhibitory effects. Only pure additive inhibitory effects were, however, obtained during simultaneous treatment of leukocytes with theophylline and histamine .Subsidiary of AB Astra, Sweden.     

Inhibition of allergic and nonallergic leukotriene C4 formation and histamine secretion by azelastine: implication for its mechanism of action

Azelastine, an orally effective antiasthmatic agent, has been reported to inhibit antihistamine-resistant, leukotriene-mediated allergic bronchoconstriction in guinea pigs. This suggests that azelastine might act through inhibition of leukotriene (LT) C4/D4 synthesis. We have examined the effect of azelastine on allergic and nonallergic histamine secretion and LTC4 formation. Azelastine and the known 5-lipoxygenase inhibitors, nordihydroguaiaretic acid and AA-861, exerted concentration-dependent inhibition of allergic LTC4 formation in chopped lung tissue from actively sensitized guinea pigs and calcium ionophore A23187-stimulated LTC4 synthesis in mixed peritoneal cells from rats. Azelastine also produced concentration-dependent inhibition of allergic and nonallergic histamine secretion from rat peritoneal mast cells. The ability of azelastine to inhibit allergic and nonallergic histamine secretion and LTC4 generation may contribute to its mode of action and its therapeutic efficacy.     

Impaired basophil histamine release from allergic patients

A few patients (6–7%) with a verified type I allergic reaction do not respond with histamine release after challenge of their basophils with specific antigen (non-responding basophils from allergic patients). Sera from these non-responding patients were used for passive sensitization of responding cells from healthy controls. When these sensitized cells were challenged with specific antigen, histamine release was observed indicating that the non-responding allergic patients have circulating antigen-specific IgE capable of binding to Fc-receptors on the basophils. These findings suggest the possibility that non-responding basophils have impaired cell functions. We therefore examined the influence of enhanced IgE receptor stimulation on histamine release in non-responding basophils. This was made by stimulating protein kinase C activity by a phorbol ester (phorbol 12-myristate 13-acetate). When the non-responding cells were incubated with the phorbol ester and challenged with either anti-IgE or specific antigen, the cells released histamine.These findings support the hypothesis that the unresponsiveness of basophils in some allergic patients is associated with impaired IgE receptor complex activation or subcellular functioning and not with a lack of cell-bound IgE.     

Inhibition of reagin-mediated PCA reactions in monkeys and histamine release from human leukocytes by human IgG4 subclass.

Human myeloma proteins of IgG4 subclass in contrast to myeloma proteins IgG1, IgG2 and IgG3, were capable of blocking PCA reactions in monkeys mediated by human reaginic antibodies of IgE class. In addition to IgE, IgG4 myeloma protein was also capable of sensitizing leukocytes from normal individuals and gave histamine release (HR) upon challenge with anti-human IgG4. Leukocytes from 11 allergic individuals and from 9 normal subjects sensitized with the serum of allergic patients, were capable of releasing histamine with anti-human IgG4, anti-human IgE, and the specific allergen. No response was obtained with anti-human IgG1 and IgG3 sera. Leukocytes from the normal individuals released histamine from 3 to 20% with anti-human IgG4 and from 6 to 30% with anti-human IgE. Moreover, normal leukocytes sensitized with IgG4 myeloma protein or a serum of an allergic patient heated at 56 degrees C for 2 h, released a significant amount of histamine on challenge with anti-human IgG4 whereas no response was obtained with anti-human IgE. The biological role of human IgG4 in immediate hypersensitivity reactions is discussed in relation to human IgE.     

Histamine release from human peripheral blood leukocytes analyzed by histamine radioimmunoassay

Histamine release from human peripheral blood leukocytes, afterin vitro challenge with allergen extracts, is usually measured by fluorometry. In the present study we compared the results of the automated fluorometric histamine assay (Siraganian) with those of the Immunotech histamine radioimmunoassay. Histamine release dose — response curves obtained after histamine measurement by both methods were superimposable.     

Cytokine-induced release of histamine from basophil leukocytes from AIDS patients

Cytokine-induced histamine release from basophil leukocytes was examined in cell suspension from AIDS patients and compared with healthy controls. Cells from approximately half of the AIDS patients, in contrast to none from the control group, showed histamine release after stimulation with interleukin-4 (IL-4), tumor necrosis factor alpha (TNF alpha), lymphotoxin (LT) and interferon gamma (IFN gamma). These cytokines seem to induce histamine release from cells from AIDS patients by interaction with the cell surface immunoglobulins, since removal of the immunoglobulins prior to the exposure of the cytokines completely abolished the response to the cytokines. IL-1 alpha, IL-1 beta, IL-3, colony stimulating factor (CSF) and granulocyte-macrophage-CSF (GM-CSF) caused significant histamine release from cells from a similar number of AIDS patients and controls.     

Inhibition of lysosomal enzyme release from rat leukocytes by auranofin

Auranofin (SK&F D-39162), a new antiarthritic gold compound reported to be orally effective in animal (adjuvant rat) and human (rheumatoid) arthritic conditions, is a potent in vitro inhibitor of the release of lysosomal enzymes from phagocytizing rat leukocytes. Auranofin, at micromolar concentrations (1–10M), produced a dose-dependent reduction in extra-cellular levels of lysosomal enzyme markers (-glucuronidase and lysozyme) which are selectively released from rat leukocytes during phagocytosis of zymosan particles. The reduction in extracellular levels of lysosomal enzymes appears to be caused by inhibition of their selective cellular release, since effective concentrations of auranofin did not produce leukocyte cytotoxicity or inhibition of cell-free lysosomal enzyme activity. Morphologic and biochemical evidence indicated that auranofin also interferes with phagocytosis of zymosan particles. The potent in vitro activity of auranofin appears to result from its unique gold complex, since neither structurally related nongold compounds nor clinically used gold compounds (gold sodium thiomalate and gold thioglucose) were potent inhibitors of lysosomal enzyme release. The results of this investigation suggest that the antiarthritic activity of auranofin may be caused at least in part, by inhibition of lysosomal enzyme release and/or cellular processing of antigens.SK&F D-39162 (2,3,4,6-tetra-O-acetyl-1-thio--D-glucopyranosato-S) (triethylphosphine) gold.Presented in part at the Federation of American Societies for Experimental Biology, Atlantic City, New Jersey, April 11, 1974 (4).     

Inhibition of passive cutaneous anaphylaxis (PCA) by azelastine: dissociation of its antiallergic activities from antihistaminic and antiserotonin properties

Azelastine and methysergide injected i.v. 5 min prior to antigen challenge and disodium cromoglycate (DSCG) injected i.v. immediately before antigen challenge produced dose-dependent inhibition of IgE-mediated 72 h passive cutaneous anaphylaxis (PCA) responses with ID50S of 0.3, 0.2 and 1.0 mg/kg, respectively. Thus, azelastine is about three times as effective as DSCG (a mast cell stabilizing agent) and somewhat less active than methysergide (a specific serotonin "D" receptor antagonist). Oral administration of azelastine and other drugs 2 h prior to antigen challenge produced strong inhibitory effects on PCA. The ID50S (mg/kg) were as follows: azelastine = 1.4; astemizole = 1.6; ketotifen = 2.0; aminophylline = 4.6; and diphenhydramine = 10.9. After 4 h of oral administration, azelastine and other drugs inhibited PCA responses with the following ID50S (mg/kg): azelastine = 1.8; astemizole = 2.3; ketotifen = 2.3; and aminophylline = 12.5. Azelastine administered orally 24 h before antigen challenge was still capable of exerting significant anti-PCA activity with an ID50 of 2.6 mg/kg, whereas none of the other drugs tested produced any significant inhibitory effects on PCA. In subsequent experiments, it was established that the antiallergic and antihistaminic activities of azelastine are inseparable 2 h after oral administration (ID50 of azelastine mg/kg, p.o., 2 h: PCA = 2.6 and histamine = 3.1). However, the persistence of the oral antiallergic (anti-PCA) effects of azelastine for 24 h (ID50 = 3.7 mg/kg) does not seem to be associated with its antihistaminic or antiserotonin activities.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibition of basophil histamine release by gangliosides

Histamine release from human basophils was inhibited by preincubation of the cells with a glucolipid mixture containing sialic acid-containing gangliosides. This was true for histamine release induced by anti-IgE, Concanavalin A and the calcium ionphore A23187, whereas the release induced by S. aureus Wood 46 was not affected. It was demonstrated that the inhibitory capacity of the glucolipid mixture could be attributed to the content of gangliosides, since no inhibition was obtained with cerebrosides or with gangliosides from which sialic acid was removed. Preincubation of the cells with the glucolipid mixture increased the sialic acid content of the cells, and this increase was attributed to an insertion of gangliosides into the cell membrane. The inhibition of histamine release was abolished by increasing the calcium concentration, which substantiates our previous findings that cell membrane sialic acid in basophil leukocytes is involved in the regulation of histamine release, possibly by a modulation of the trans-membraneous calcium fluxes preceding histamine release.