Anaphylaxis In Rats Research Articles

ＹＭ‐１３６５０のＩ〜ＩＶ型アレルギー反応および抗体産生に対する作用

The effects of YM-13650 on experimental animal models of cell-mediated immune responses (type IV), antibody formation and type I to type III allergic reactions were investigated. YM-13650 in the dose range of 6.3 to 100 mg/kg, p.o., inhibited the picrylchloride-induced delayed type hypersensitivity (Pc-DTH) in mice when administered during the induction and the effector phases. The compound also inhibited the Pc-DTH enhanced by the pretreatment with cyclophosphamide, and even bilateral adrenalectomy failed to reduce the inhibitory effect of the compound on Pc-DTH in mice. YM-13650 in doses of 25 and 50 mg/kg, p.o., prolonged the survival time of allogenic skin grafts in mice. However, no significant effect was observed on hapten-specific IgE and HA antibody production and PFC formation in mice in doses up to 300 mg/kg, p.o. YM-13650 inhibited the passive Arthus reaction in guinea pigs and the reversed passive Arthus reaction in rats (type III). On the other hand, YM-13650 did not show any inhibitory effect on passive cutaneous anaphylaxis in rats (type I), Forssman shock in guinea pigs (type II) and carrageenin-induced paw edema in rats. These results indicate that YM-13650 suppresses not only cell-mediated immune responses but also type III allergic reactions without any influence on type I and type II allergic reactions as well as an acute inflammatory reaction.

柑橘類果実の薬理学的研究(第2報)ウンシュウミカン果実の抗アレルギー作用その2 フラボノイド成分について

A study was carried out to clarify the active component of green fruit of Citrus unshiu MAKOVICH on the type I allergic reaction. The flavonoid component, isolated from its methanolic extract, hesperidin, was found to inhibit histamine release from peritoneal mast cells of rats induced by compound 48/80. Forty eight-hour homologous passive cutaneous anaphylaxis (PCA) in intact rats was significantly inhibited by the oral administration of hesperidin. However, the anti-allergic action on PCA was not observed in adrenalectomized rats. These results suggests that hesperidin is an effective component of the fruit of Citrus unshiu with the anti-allergic action against the type I reaction.

Molecular cloning of grass pollen allergens: progress and perspectives

In this report the advances made in recent years as a result of application of recombinant DNA and allied technologies in relation to isolation, molecular characterization and synthesis of principal allergens—with particular emphasis on Kentucky Bluegrass (KBG,Poa pratensis) pollen allergens—has been reviewed. Screening of a cDNA library complementary to KBG pollen mRNA led to isolation of several cDNA clones. Hybridization studies using the labelled cDNAs as probes suggested the existence of at least three major classes of clones. The nucleotide sequences of several cDNAs encoding a major group of allergenic proteins were determined. Moreover, one cDNA clone, KBG7.2 was inserted in an expression plasmid to achieve high-level expression of the corresponding peptide. The recombinant peptide was shown to be comparable to its natural counterpart with regard to its ability to bind to human IgE and IgG antibodies of allergic patients, and to elicit passive cutaneous anaphylaxis (PCA) in rats. Furthermore, KBG 7.2 encoded peptide was shown to possess at least two IgE binding epitopes. About four T cell epitopes were predicted on this peptide. The results of these studies suggest that the recombinant allergens and/or allergenic peptides may be of diagnostic and therapeutic value.

Hemodynamic changes associated with anaphylaxis in parasite-sensitized rats

Thirty days after infection with the parasite Nippostrongylus brasiliensis, microcirculatory changes associated with worm allergen-induced anaphylactic shock in pentobarbital-anesthetized rats were studied. Allergen challenge decreased heart rate, mean arterial blood pressure, and cardiac output (CO) for 60 min, although CO increased markedly by 240 min. Blood flow (ml.min-1.g wet tissue-1) was reduced to most tissues, with the exception of the kidneys, hepatic arterial, heart, and brain, for 60 min. Depressed flow to the adrenals and enhanced flow to the kidneys, trachea, stomach, and mesentery were observed at 240 min. The percent distribution of cardiac output per gram wet tissue (%CO/g) increased initially to the kidneys, heart, liver, and cervical spinal cord, decreased to the spleen and cecum, and remained unchanged to other gastrointestinal tissues. The %CO/g to most tissues returned to normal by 60 min but decreased to the adrenals at 240 min. The intestine is an important organ affected by anaphylaxis in the rat, and the liver may participate in alleviating anaphylaxis.

大豆粕乾留タール（Ｇｌｙｔｅｅｒ）の抗炎症作用 （第３報） Ｉ‐ＩＶ型アレルギー反応に対する作用

Effects of Glyteer (GL, 5%) on Type I-Type IV allergic reactions were investigated by its topical application to rats and mice, and the effects were compared with those of betamethasone 17-valerate (BV, 0.12%), indomethacin (ID, 1%), and bufexamac (BM, 5%), which were all prepared with the same ointment base. Type I: GL showed inhibitory effects on the 48 hr homologous passive cutaneous anaphylaxis (PCA) in rats. The inhibitory activity of GL on the PCA had the same potency as that of BV (0.12%). GL also inhibited the degranulation of mast cells induced by PCA. Type II: GL did not exert an inhibitory effect on the reversed cutaneous anaphylaxis (RCA) in rats, but BM, ID and BV had an inhibitory activity on the RCA. Type III: BV markedly inhibited the direct passive Arthus reaction in rats. On the other hand, GL, BM and ID had not an inhibitory activity on it. Type IV: GL (0.2, 1 and 5%) showed a concentration-dependent inhibition on the delayed-type hypersensitivity response induced by oxazolone in mice, and the activity was stronger than those of ID and BM. From these results, it is suggested that GL applied externally possesses a potent effect as an anti-allergic drug on Type I and Type IV allergic reactions.

非ステロイド性抗炎症薬Ａｌｍｉｎｏｐｒｏｆｅｎの抗アレルギー作用

The effects of alminoprofen, a nonsteroidal anti-inflammatory agent, on the experimental animal models of allergic reactions were examined and compared with those of ibuprofen. Alminoprofen at 30 mg/kg given intraduodenally significantly suppressed passive anaphylactic bronchoconstrictions, while ibuprofen did not at the same doses. In vitro studies revealed that alminoprofen, in contrast to ibuprofen, exerted an inhibitory effect on arachidonate 5-lipoxygenase activity which initiates the bio-synthesis of leukotrienes. Alminoprofen inhibited arachidonic acid-induced ear edema in mice and homologous passive cutaneous anaphylaxis in rats at high doses, while ibuprofen did not at the high doses. From its characteristic feature of inhibitory effects on 5-lipoxygenase activity and the experimental model of type I allergic reaction, it is suggested that alminoprofen is a new type of nonsteroidal anti-inflammatory agent.

Antiallergic and Anti-Inflammatory Action of Tioxamast in Rats I.

Tioxamast (F 1865) is an antiallergic drug that, administered systemically, reduces anaphylaxis in various models in rats. This action is due mainly to the inhibition of the synthesis and release of certain mediators. Orally or intraduodenally administered tioxamast inhibits IgE-dependent passive cutaneous anaphylaxis (ED50 = 0.8 mg/kg), IgE-dependent passive pulmonary anaphylaxis (ED50 = 0.5 mg/kg), and IgG-dependent passive cutaneous anaphylaxis (ED50 = 0.6 mg/kg). It has little or not effect on the increase of cutaneous capillary permeability induced by various mediators. In IgE-dependent passive peritoneal anaphylaxis in rats, tioxamast reduces the release of histamine (IC50 = 0.024 micrograms/ml) and of beta-glucuronidase (IC50 = 0.102 micrograms/ml). Also, histamine release is inhibited in IgG-dependent peritoneal anaphylaxis (IC50 = 0.103 micrograms/ml). The antiallergic compound has less effect on the release of histamine induced by the compound 48/80 in the peritoneal cavity of rats (IC50 = 1.67 micrograms/ml). Tioxamast inhibits the synthesis in vitro of leukotriene B4 (LTB4) by peritoneal neutrophils from rats stimulated by A23187 (IC50 = 8.88 micrograms/ml). At higher tioxamast concentrations, metabolites of the cyclo-oxygenase pathway are inhibited at concentrations of the same order of magnitude as those that inhibit Naja naja phospholipase A2 (IC50 = 144 micrograms/ml). Tioxamast also reduces the production of free radicals by leukocytes from the pleural cavity of rats which had phagocytosed opsonized zymosan (IC50 = 5.21 micrograms/ml).

Potent Inhibitory Activity of HSR-6071, a New Antiallergic Agent, on Passive Cutaneous Anaphylaxis (PCA)

Antiallergic effects of 6-(1 -pyrrolidinyl)-N-(1H-tetrazol-5-yl)-2-pyraz-inecarboxamide (HSR-6071), a newly synthesized agent, were investigated. The 48-hr homologous passive cutaneous anaphylaxis (PCA) in rats was inhibited in a dose-dependent manner by i.v. and p.o. administration of the agent (ED50=0.0096 mg/kg and 0.18 mg/kg, respectively). The IgE-mediated histamine release from rat peritoneal exudate cells was inhibited by HSR-6071, with an IC50 of 4.6×10−10 M. Regarding the non-immunological histamine release, HSR-6071 inhibited compound 48/80-induced, but not A23187-induced and spontaneous histamine release. On the other hand, an increase in vascular permeability induced by histamine, serotonin and bradykinin was unaffected by HSR-6071 in doses sufficient to inhibit PCA. In addition, the contractile responses of isolated guinea pig ileum to histamine, acetylcholine and serotonin were also unaffected by the agent even in a high concentration of 10−4 M. These results indicate that HSR-6071 possesses a potent antiallergic activity and that the inhibition of PCA by HSR-6071 may be due to the suppression of chemical mediators release from mast cells.

Potent inhibitory activity of HSR-6071, a new antiallergic agent, on Passive Cutaneous Anaphylaxis(PCA).

Antiallergic effects of 6-(1-pyrrolidinyl)-N-(1H-tetrazol-5-yl)-2-pyrazinecarboxamide (HSR-6071), a newly synthesized agent, were investigated. The 48-hr homologous passive cutaneous anaphylaxis (PCA) in rats was inhibited in a dose-dependent manner by i.v. and p.o. administration of the agent (ED50 = 0.0096 mg/kg and 0.18 mg/kg, respectively). The IgE-mediated histamine release from rat peritoneal exudate cells was inhibited by HSR-6071, with an IC50 of 4.6 x 10(-10) M. Regarding the non-immunological histamine release, HSR-6071 inhibited compound 48/80-induced, but not A23187-induced and spontaneous histamine release. On the other hand, an increase in vascular permeability induced by histamine, serotonin and bradykinin was unaffected by HSR-6071 in doses sufficient to inhibit PCA. In addition, the contractile responses of isolated guinea pig ileum to histamine, acetylcholine and serotonin were also unaffected by the agent even in a high concentration of 10(-4) M. These results indicate that HSR-6071 possesses a potent antiallergic activity and that the inhibition of PCA by HSR-6071 may be due to the suppression of chemical mediators release from mast cells.

Ｔａｚａｎｏｌａｓｔ （ＷＰ‐８３３） 代謝物の抗アレルギー作用

WP-833 is an orally active new antiallergic drug. The effects of WP-833 metabolites on experimental allergic reactions were investigated in various animal models. MTCC [3'-(1H-tetrazol-5-yl) oxanilic acid], the main metabolite of WP-833, dose-dependently inhibited 48-hr homologous passive cutaneous anaphylaxis (PCA) in rats and 8-day PCA in guinea pigs after intravenous administration. The inhibitory effect of MTCC on PCA was nearly equal to that of WP-833. MTCC also inhibited 48hr PCA in rats after the peroral administration, although its inhibitory effect was about 6 times weaker than that of WP-833. MTCC also showed a dose-dependent inhibition on the release of histamine or slow reacting substance of anaphylaxis (SRS-A) from passively sensitized rat peritoneal mast cells and guinea pig lung fragments in vitro with a similar potency to WP-833. MTA [3-(1H-tetrazol-5-yl) aniline] and MTAA [3'-(1H-tetrazol-5-yl) acetanilide], other metabolites of WP-833, inhibited PCA and anaphylactic histamine release from rat peritoneal mast cells at considerably higher doses than WP-833, and they showed no effects on the release of histamine and SRS-A from guinea pig lung fragments. Moreover, only small amounts of MTA and MTAA were detected in the blood after the oral administration of WP-833. From these results, it was suggested that MTCC may be the main active metabolite of WP-833.

Studies on antiallergic agents. I. Synthesis and antiallergic activity of novel pyrazine derivatives.

Various pyrazine derivatives were synthesized and their antiallergic activity was examined. The inhibitory activity on allergic histamine release of the compounds bearing a 5-tetrazolyl group was more potent than that of the corresponding carboxyl derivatives. The introduction of -CONH- or -NHCO- between the pyrazine ring and the 5-tetrazolyl group as a spacer greatly enhanced the activity. N-(1H-Tetrazol-5-yl)-2-pyrazinecarboxamide (I-3) was estimated to exhibit nearly the same potency as disodium cromoglycate (DSCG). The structure-activity relationship among various derivatives modified by introducing some substituents onto the 3-, 5- or 6-position of the pyrazine ring of I-3 was investigated. The activity remained unchanged or was reduced when such substituents as methyl, chloro, methoxy, methylamino and dimethylamino were introduced at the 3- or 5-position. In contrast, 6-substitution with various alkylamino groups more or less increased the activity. Among them, the 6-dimethylamino (I-17c) and 6-(1-pyrrolidinyl) (I-34) derivative were proved to be most potent. The IC50 values (concentration which produces 50% inhibition of the allergic histamine release) of I-17c and I-34 were determined to be 4.7 x 10(-10) and 4.6 x 10(-10) M, respectively. These two compounds produced a potent inhibitory activity on passive cutaneous anaphylaxis (PCA) in rat, not only by the intravenous route (ED50 = 0.0096 mg/kg for both compounds) but also by the oral route (ED50 = 0.19 and 0.18 mg/kg, respectively). On the other hand, when the pyrazine ring of some representative compounds was replaced with a pyridine ring, the inhibitory activity on histamine release was significantly reduced.

Intestinal anaphylaxis in the rat: Mediators responsible for the ion transport abnormalities

Antigen challenge of jejunal epithelium from rats sensitized to egg albumin induces an active Cl- secretory process secondary to release of mucosal mast cell mediators. The present study was designed to define the relative role of these mast cell mediators and the enteric nervous system in the transport abnormalities associated with intestinal anaphylaxis. Net ion transport of stripped jejunal tissue from sensitized and sham-treated animals was studied in Ussing chambers. The Cl- secretory response induced by egg albumin during intestinal anaphylaxis was similar to that after addition of 5-hydroxytryptamine (5-HT), histamine, and prostaglandins D2 and E2 to jejunal tissue. Cinanserin, a 5-HT2-receptor antagonist, virtually abolished the response to 5-HT and totally abolished the response to egg albumin. Methysergide, a 5-HT1-receptor antagonist had no effect on either response. Indomethacin, an inhibitor of prostaglandin synthesis, significantly inhibited the 5-HT and egg albumin response. Diphenhydramine, an H1-receptor antagonist and cimetidine, an H2-receptor antagonist both significantly inhibited the histamine response but neither altered the response to egg albumin. Atropine, an anticholinergic, and tetrodotoxin, a nerve blocker, did not inhibit the antigen induced anaphylactic response. These results indicate that 5-HT, acting through 5-HT2 receptors is largely responsible for the transport abnormalities seen in intestinal anaphylaxis induced by egg albumin while prostaglandins appear to play a partial role. The findings do not support a role for the enteric nervous system for the egg albumin induced changes in Cl- secretion.

Synthesis and pharmacological evaluation of procaterol derivatives having a tert-amino group.

A series of procaterol derivatives having a tert-amino group was synthesized. Among them, a morpholino derivative (4a, R1 = H, NR2R3 = morpholino) showed beta-selective and rather potent adrenoceptor stimulant activities in an in vivo assay using anesthetized dogs. On the other hand, a morpholinopropanol analogue (4j, R1 = CH3, NR2R3 = morpholino) showed 400 times less potent bronchodilator activity than that of 4a. Some of the compounds showed weak bronchodilator activities and weak effects on the heart. It seems that steric hindrance around the nitrogen atom of catecholamines has a significant influence on beta-adrenoceptor stimulant activities. Compound 4a also showed anti-allergic action estimated in terms of the inhibition of homologous passive cutaneous anaphylaxis in rats.

Inhibitory Effects of β-Adrenergic Stimulants on Increased Vascular Permeability Caused by Passive Cutaneous Anaphylaxis, Allergic Mediators, and Mediator Releasers in Rats

The effects of isoproterenol, salbutamol, theophylline, and forskolin on IgE antibody-mediated homologous passive cutaneous anaphylaxis (PCA) and on skin reactions caused by allergic mediators and mediator releasers were investigated in rats. Isoproterenol and salbutamol dose-dependently inhibited the PCA and skin reactions caused by histamine, serotonin, and leukotriene C4 (LTC4), elicited at the same time in the same rat. These agents also dose-dependently inhibited the skin reactions caused by platelet-activating factor (PAF), leukotriene D4 (LTD4), compound 48/80 (48/80), and calcium ionophore A23187 (A23187). Propranolol overcame the inhibitory effect of isoproterenol on PCA and skin reactions in a dose-dependent manner. Theophylline and forskolin showed similar effects as the beta-adrenergic stimulants. These results indicate that beta-adrenergic stimulants inhibit the increased vascular permeability caused by allergic mediators, and suggest that this activity of beta-adrenergic stimulants might play an important role in their antiallergic actions. Inhibition of increased vascular permeability might be mediated via beta-receptors and may be related to the increase in intracellular cyclic AMP levels.

柑橘類果実の薬理学的研究(第1報)ウンシュウミカン果実の抗アレルギー作用 その1

A study was carried out to examine the effect of 50% ethanolic extract from green fruit of Citrus unshiu Markovich on type I, II and IV allergic reactions. Forty eight-hour homologous passive cutaneous anaphylaxis (PCA) in rats as a typical model of the type I reaction was significantly inhibited by the oral administration of the extract. Complement-dependent cytolysis of sheep red blood cell as a typical model of the type II reaction was significantly inhibited by incubation with it. Contact dermatitis in mice as a model of the type IV reaction caused by picryl chloride or sheep red blood cell was significantly inhibited by the oral application of it. The anti-allergic actions decreased with the growth of fruit of Citrus unshiu. These results suggested that the 50% ethanolic extract from green fruit of Citrus unshiu have anti-allergic actions against the type I, II and IV reactions.

Lack of involvement of leukotriene and platelet activating factor in passive cutaneous anaphylaxis in rats.

Possible chemical mediators contributing to 48 hour passive cutaneous anaphylaxis (PCA) in rats were investigated. Forty-eight hour PCA was inhibited considerably by mepyramine and methysergide given intravenously, a finding suggestive of a major role for histamine and serotonin in the reaction. AA-861, a selective 5-lipoxygenase inhibitor did not inhibit the PCA, and leukotriene (LT)D4 or LTE4 and the combination with prostaglandin (PG)E2 had no significant skin reaction. In addition, only small amounts of slow reacting substance of anaphylaxis (SRS-A) were detected in skin fragments, in vitro. Although CV-3988, a selective platelet activating factor (PAF) antagonist, dose-dependently inhibited the PAF-induced skin reaction, the PCA was not affected by treatment with this compound. Indomethacin also had no inhibitory activity on PCA. Thus, sulfidopeptide LTs, PAF and arachidonate cyclooxygenase metabolites probably do not contribute to PCA, at least in rats.

Mechanisms of the antiallergic action of N-methylmequitazine (LG 30435).

LG 30435 (N-methylmequitazine) was assayed in passive lung (PLA) and cutaneous (PCA) anaphylaxis in guinea-pigs and rats. At doses from 0.3 to 3 mumol kg-1 i.v., it produced a dose-dependent inhibition of guinea-pig PLA and of rat PCA and PLA, while the parent compound was ineffective or poorly effective up to 3 mumol kg-1. An attempt was made to elucidate the mechanism of LG 30435's action in these anaphylactic models, by means of various antagonists. It was tentatively concluded that different mechanisms are involved in the protective action of LG 30435 in each of the three models: histamine antagonism, possibly accompanied by an inhibition of the effects of peptido-leukotrienes in guinea-pig PLA; histamine antagonism in rat PCA and 5-hydroxytryptamine antagonism in rat PLA, possibly accompanied by a mast-cell stabilizing action in both cases. LG 30435 is devoid of smooth muscle relaxant effects on the airways and its demonstrated anticholinergic and anti-PAF effects do not appear to be involved in its antiallergic action.

Antiallergic effects of mequitazine. 2. In vivo experiments

Antiallergic effects of mequitazine were investigated in vivo and compared with those of ketotifen and disodium cromoglycate (DSCG). The results obtained were as follows: 1) Mequitazine in doses of 2 and 5 mg/kg given, p.o., 1 hr prior to antigen challenge inhibited dose-dependently the 48-hr passive cutaneous anaphylaxis in rats. Five mg/kg of mequitazine showed almost the same extent of inhibitory activity as that of 1 mg/kg of ketotifen. An i.v. administration of 1 mg/kg DSCG 1 min before antigen challenge also showed a marked inhibition. 2) The experimental asthma induced by challenge with an i.v. injection of antigen in passively sensitized guinea pigs was fairly inhibited by the pretreatment with 5 mg/kg of mequitazine administered p.o., although 2 mg/kg of this drug showed only a slight inhibition. 3) The experimental asthma induced by aerosolized antigen was also fairly inhibited by the pretreatment with 5 mg/kg of mequitazine given p.o.

Sensitivity of passive cutaneous anaphylaxis in rats. II. Suppression of passive cutaneous anaphylactic reactions in rats infected with Nippostrongylus brasiliensis.

The sensitivity of passive cutaneous anaphylactic (PCA) reaction was examined in rats infected with the nematode parasite, Nippostrongylus brasiliensis (Nb). PCA reactions with anti-DNP IgE antibody were remarkably suppressed by Nb infection already 10 days after infection and persisted for at least 28 weeks. 10 days after infection, no anti-Nb IgE antibody was present in the sera of infected rats, whereas 18 or 28 weeks after infection, circulating anti-Nb IgE antibody was present. It was concluded that two different mechanisms might explain the suppression. 10 days after infection, nonspecific IgE induced by potentiated IgE production inhibited passive sensitization with anti-DNP IgE antibody by occupying the IgE receptors on mast cells, whereas 18 or 28 weeks after infection, the mast cell IgE receptors were occupied by IgE antibody specific for the Nb antigen. The reactivity through non-immunological processes (skin sensitivity to compound 48/80) was not modified by Nb infection at any time after infection.

Attenuation of rat conjunctival response by repeated hapten applications.

Attenuation of the rat conjunctival response by repeated topical challenge with dinitrophenyl (DNP) hapten was demonstrated in our study. Adult rats were immunized by intraperitoneal injections of dinitrophenylated Ascaris suum extract (DNP-Asc) and alum. Serum levels of anti-DNP homocytotropic antibody were determined by passive cutaneous anaphylaxis in rats prepared with antibody 48 hours earlier. In other animals, topical challenge was performed by applying N,N'-di-2,4-DNP-L-lysine (di-DNP-lysine) in phosphate-buffered saline (PBS) to one eye; PBS alone was applied to the fellow eye. The degree of conjunctival reaction was assessed clinically, and ocular tissues were processed for histological evaluation. The intensity of the conjunctival reaction and extent of mast cell degranulation were significantly greater after one challenge with di-DNP-lysine than after multiple challenges. In the multiple-challenge group, the contralateral eye remained responsive to a single challenge with di-DNP-lysine. These results may have implications for therapeutic interventions in ocular anaphylaxis.