Search

Background The expression and functional role of CysLT 2 receptors in asthma have not been clarified. In this study, we evaluated CysLT 2 receptors expression, and effects of CysLT 2 -and CysLT 1/2 -receptor antagonists on antigen-induced bronchoconstriction using isolated lung tissues from both asthma and non-asthma subjects. Methods CysLT 1 and CysLT 2 receptors expression in asthma and non-asthma lung tissue preparations was examined in immunohistochemistry experiments, and their functional roles in antigen-induced bronchoconstriction were assessed using ONO-6950, a dual CysLT 1/2 -receptor antagonist, montelukast, a CysLT 1 receptor antagonist, and BayCysLT 2 RA, a CysLT 2 receptor-specific antagonist. Results CysLT 1 receptors were expressed on the bronchial smooth muscle and epithelium, and on alveolar leukocytes in 5 in 5 non-asthma subjects and 2 in 2 asthma subjects. On the other hand, although degrees of CysLT 2 receptors expression were variable among the 5 non-asthma subjects, the expression in the asthma lung was detected on bronchial smooth muscle, epithelium and alveolar leukocytes in 2 in 2 asthma subjects. In the non-asthma specimens, antagonism of CysLT 2 receptors did not affect antigen-induced bronchial contractions, even after pretreatment with the CysLT 1 -receptor specific antagonist, montelukast. However, in the bronchus isolated from one of the 2 asthma subjects, antagonism of CysLT 2 receptors suppressed contractions, and dual antagonism of CysLT 1 and CysLT 2 receptors resulted in additive inhibitory effect on anaphylactic contractions. Conclusions CysLT 2 receptors were expressed in lung specimens isolated from asthma subjects. Activation of CysLT 2 receptors may contribute to antigen-induced bronchoconstriction in certain asthma population.

Hairless mice fed a special diet, HR-AD, develop atopic dermatitis (AD)-like skin inflammation with skin barrier defects and itch-related scratching; however, the ingredient(s) causing the dermatitis remains unclear. In this study, we examined whether deficiency of certain polyunsaturated fatty acids (PUFAs) is involved in HR-AD-induced AD. High-purity PUFAs were given to HR-AD-fed mice by dietary supplementation or gavage. Fatty acid levels in the serum and skin were determined by using gas chromatography-mass spectrometry. In serum from HR-AD-fed mice, linoleic acid (LA, 18:2n-6) and α-linolenic acid (ALA, 18:3n-3), as well as their metabolites, were markedly decreased. When mice were fed HR-AD supplemented with LA or ALA in an amount equal to that contained in a normal diet, the development of AD-like symptoms was completely prevented by supplementation with LA but not with ALA. Relatively high dose of ALA slightly alleviated skin barrier defects, but did neither itch-related scratching nor skin inflammation. On the other hand, gavage administration of LA metabolites, such as γ-linolenic acid and arachidonic acid (AA), significantly ameliorated established dermatitis without increasing LA in the serum and skin. Moreover, AA-induced amelioration of dermatitis was not affected by pharmacological blockade of 5-lipoxygenase (5-LOX) and cyclooxygenase (COX), suggesting no involvement of 5-LOX- or COX-mediated AA metabolites in the amelioration. In conclusion, our results indicate that deficiency of n-6 PUFAs is mainly responsible for AD-like symptoms by HR-AD feeding. Thus, this model could be useful for studying the pathomechanisms associated with deficiency of n-6 PUFAs in AD.

In atopic dermatitis (AD), severe itching and scratching not only impacts the patient's quality of life, but also exacerbates the disease; however, the precise underlying mechanisms of itching in AD remain elusive. We have previously reported a special diet-induced, mouse model of AD. This article reviews the characteristics of the model, and the chronological and pharmacological analyses of the pathology. HR-1 hairless mice fed a commercial special diet, HR-AD, but not a normal diet, showed scaly dry red skin resembling AD. To evaluate the degree of itching, spontaneous scratching was analyzed. As a result, scratching with long duration was observed in HR-AD-fed mice, as confirmed by prolongation of duration of one scratching bout. Chronological analysis of the pathologies in HR-AD-fed mice revealed that the prolonged scratching was coincident with the development of skin barrier dysfunction, while skin inflammation with recruitment of inflammatory cells, such as CD4+ T cells, eosinophils, and mast cells, and serum IgE elevations apparently followed the prolonged scratching. Epidermal nerve sprouting was detected almost coincidently with the prolonged scratching. Pharmacological analysis of the scratching behavior indicated that the prolonged scratching was suppressed by a μ-opioid receptor antagonist, but not by antihistamine, similar to AD patients. Although treatment with corticosteroid suppressed the skin inflammation, it was not effective against the prolonged scratching, the dry skin symptoms, and the epidermal nerve sprouting. Petrolatum ointment inhibited the prolonged scratching during the amelioration of skin barrier dysfunction. Furthermore, the barrier disrupted-dry skin was alleviated by oral administration of essential fatty acid, linoleic acid (LA), suggesting that LA deficiency appears to be responsible for the symptoms. Taken together, these findings suggest that skin barrier dysfunction is closely associated with the development of AD-like pruritus in this model.

Background Japanese cedar pollen-induced allergic rhinitis in a guinea pig model clearly induced not only sneezing but also biphasic nasal blockage. To date, there have only been a few reports on models of murine allergic rhinitis which clearly show nasal blockage. Therefore, in order to try and develop such a model, we administered multiple dosages of intranasal pollen or purified antigen protein Cry j 1. Methods B10.S mice were sensitized by intranasal instillations of either pollen extract or Cry j 1 twice a day for 7 days, which was adsorbed on Al(OH) 3 . Subsequently, once a week, the mice were given multiple intranasal instillation challenges of either the pollen suspension or Cry j 1 and the frequency of sneezing was observed after respective challenges were made. Specific airway resistance (sRaw) was measured as an indicator for nasal blockage. Cry j 1-specific IgE levels were measured using an enzyme-linked immunosorbent assay. Results The serum Cry j 1-specific IgE level showed clear elevation only in the group sensitized by Cry j 1 + Al(OH) 3 and then challenged by Cry j 1. No elevations were seen in the groups sensitized by pollen extract + Al (OH) 3 followed by a pollen suspension challenge. There was an immediate increase in sneezing after challenges in all of the sensitized-challenged groups. Nevertheless, no increases in sRaw in any of the groups were detected at any of the time points during the 8 hours following the challenges. Conclusions Cry j 1 may be more effective than crude antigens for efficient sensitization/challenge in mice. No increase in sRaw occurred, even in mice that possessed high amounts of Cry j 1-specific IgE and that exhibited sneezing.

BACKGROUND Although it has been suggested that the use of tachykinin receptor antagonists might prove to be an effective treatment for allergic rhinitis (AR), they are not used clinically. Therefore, we decided to examine the effects of tachykinin receptor antagonists on AR symptoms in an appropriate experimental model. OBJECTIVE To evaluate newly developed tachykinin receptor antagonists in a Japanese cedar pollen-induced AR model and to determine their effect on allergen-induced sneezing, nasal blockage, and nasal hyperresponsiveness (NHR). METHODS Sensitized guinea-pigs were challenged by forced inhalation of pollen once every week. Sneezing and nasal blockage were observed after pollen challenges. NHR (nasal blockage) to an intranasal application of leukotriene D(4) was assessed 2 days after an antigen challenge. We also evaluated whether intranasal dosing with a tachykinin causes NHR. NK(1) and NK(2) receptor antagonists were administered before an intranasal treatment with antigen or tachykinin. Amounts of tachykinins present in nasal cavity lavage fluid were measured by an enzyme immunoassay. RESULTS Although an NK(1) and NK(2) receptor dual antagonist showed no effect on pollen-induced sneezing and biphasic nasal blockage, it did completely suppress the development of NHR. Experiments using specific NK(1) or NK(2) receptor antagonists revealed that NK(2) receptor activation was preferentially involved in the development of hyperresponsiveness. Increases in the levels of substance P (SP) and neurokinin A (NKA) in the nasal tissue were noted 20 min-1 h after the challenge. Intranasal instillation of either SP or NKA-induced NHR, which was almost completely inhibited by NK(2) receptor antagonists and partially inhibited by NK(1) receptor antagonists. CONCLUSIONS SP and NKA, which are released early after the challenge, mediate the development of NHR by preferentially activating NK(2) receptors. Therefore, NK(2) receptor antagonists might prove to be effective treatment of AR.

Exposure of animals to cigarette smoke for longer than 3 months leads to the development of chronic obstructive pulmonary disease (COPD) showing pulmonary emphysema. We attempted to create a COPD model with emphysema that could be established in a shorter period of time. Guinea pigs were intratracheally treated once a day on days 0-3, 5-8, 10-13 and 15-18 with a cigarette smoke solution (CSS), which was prepared by bubbling a stream of smoke into saline. Additionally, lipopolysaccharide (LPS) was administered intratracheally as an exacerbation factor on days 4, 9 and 14. By day 19, there was a gradual elevation of specific airway resistance (sRaw). In addition, both residual volume and functional residual capacity were found to be significantly higher on day 19. In the lungs, there was a marked increase in leukocytes, especially neutrophils. Histologically, we observed epithelial hyperplasia and emphysema. On the other hand, daily oral administration of theophylline during the administration of CSS and LPS suppressed the sRaw increase and the epithelial hyperplasia, but not other functional structural changes. In conclusion, we established an experimental COPD model in guinea pigs by using intratracheal instillations of CSS and LPS over a considerably shorter term than has been reported for other models.

KP-496 is a novel dual antagonist for leukotriene (LT) D(4) and thromboxane (TX) A(2) receptors. We investigated effects of KP-496 on antigeninduced nasal blockage in 2 guinea pig models of allergic rhinitis.Male Hartley guinea pigs were used.Animals were actively sensitized with ovalbumin (OVA) or Japanese cedar pollen, and were then repeatedly challenged with OVA or pollen, respectively. KP-496 (0.003 %-0.05 %) was intranasally administered 0.5 or 1 h before and 2 h after an antigen challenge.As an indicator of nasal blockage, specific airway resistance was measured using a double-flow plethysmograph system. Statistical analyses were performed with Dunnett's test (OVA model) or t-test (pollen model).Although early phase response was not affected by even a high dose (0.03 %) of KP-496, late phase nasal blockage (1.68 +/- 0.26) was inhibited by 0.01 % (0.87 +/- 0.19; p0.05) and 0.03 % (0.44 +/- 0.12; p0.01) of KP-496 in the OVA model. On the other hand, both early (5.60 +/- 0.77) and late phase responses (7.90 +/- 1.70) were inhibited by 0.05 % KP-496 to 2.68 +/- 0.84 (p0.05) and 2.71 +/- 0.83 (p0.05), respectively, in the pollen model, in which nasal hyperresponsiveness had been acquired by multiple challenges.KP-496 may be clinically effective for nasal blockage in allergic rhinitis.

Background: There is little information about the involvement of galectin-9 (Gal-9) in allergic inflammation. Thus, we investigated the role of Gal-9 in asthma model guinea pigs. Methods: Airway resistance (Raw) was measured using a double-flow plethysmograph system. Gal-9 expression in the lung was assessed by Western blot and immunohistochemistry. Eosinophil chemotactic activity was evaluated in a chamber containing a polyvinylpyrolidone-free membrane. Cell apoptosis was analyzed on a flowcytometry with propidium iodide. Results: In cloning guinea pig Gal-9 we identified three isoforms that differ only in the length of their linker peptides, just as with human Gal-9. Guinea pig Gal-9 was found to be a chemoattractant for eosinophils and to promote induction of apoptosis in sensitized but not non-sensitized T lymphocytes. In allergic airway hypersensitivity model, a low level of Gal-9 expression was observed in the nonsensitized/nonchallenged group, but upregulation was detected at 7 h after challenge and sustained up to 24 h. Such upregulation correlated with elevation of eosinophil peroxidase activity but not with increased Raw. Conclusions: The present results provide evidence that Gal-9 is not involved in airway hypersensitivity, but is partly involved in prolonged eosinophil accumulation in the lung.

Itching is the most important symptom in atopic dermatitis because the persistent scratching in response to itching aggravates the disease. However, the etiologic mechanisms of itching in atopic dermatitis remain uncertain. HR-1 hairless mice fed a special diet, HR-AD, develop atopic dermatitis-like symptoms with prolonged scratching episodes. The purpose of this study was to examine whether skin nerve fiber changes were involved in the prolonged scratching seen in this mouse model. On day 56 after the start of feeding, prolonged scratching, as well as atopic dermatitis-like skin changes, were clearly observed in HR-AD-fed mice, while no abnormal changes were observed in mice fed a normal diet. Immunohistochemical analyses of the skin using antibody to protein gene product 9.5 showed the development of numerous immunoreactive nerve fibers in the epidermis of HR-AD-fed mice. Furthermore, after cessation of HR-AD feeding, the reduction in intraepidermal nerve fibers coincided with decreased scratching. Neither the prolongation of scratching nor the increase in intraepidermal nerve fibers was affected by dexamethasone treatment. Thus, the increased number of intraepidermal nerve fibers could be involved in the aggravation of itch-related scratching observed in this model. Keywords:: itch, atopic dermatitis, nerve fiber, hairless mice, protein gene product 9.5

Dry skin/barrier dysfunction is considered to be one of the characteristic features of atopic dermatitis (AD). When HR-1 hairless mice are fed a special diet, HR-AD, dry red skin is induced. We examined whether HR-AD-fed mouse could be used as a model for AD by showing itch-associated scratching behaviour and by analysing the immunological change. HR-1 mice were fed HR-AD from 4 weeks old. HR-AD-fed mice showed severe dry skin symptoms accompanied by a decrease in dermal water content and an increase in transepidermal water loss and prolonged scratching bout duration on day 14 or 28. These symptoms became gradually worse until day 56. Marked epidermal hyperplasia and slight increase in CD4+ cells in the skin were observed from day 28. In contrast, increases in circulating T cells and serum immunoglobulin E were seen from day 41. Other skin-infiltrating inflammatory cells, such as eosinophils and mast cells, were increased on day 56 but not on day 28. Though daily oral treatment with dexamethasone reduced the increased numbers of these cells, it did not affect the dry skin symptoms or the prolonged scratching episodes. In contrast, the development of dry skin was inhibited by feeding with 10% normal diet-containing HR-AD. The skin barrier dysfunction in HR-AD-fed mice is closely associated with the development of AD-like pruritus. Changes in the immunological parameters observed may be the consequence of skin barrier dysfunction. Our findings suggest that HR-AD-fed mouse could be used as a dry skin-based experimental model for AD.

：アレルギー性鼻炎はくしゃみ，鼻汁分泌亢進および鼻閉を主症状とする典型的なI型アレルギー疾患である．くしゃみおよび鼻汁分泌は抗ヒスタミン薬で強く抑制されるが，患者にとって最も苦痛を強いる鼻閉の治療にはグルココルチコイドが繁用され，これに代わる鼻閉の治療薬が望まれる．我々はスギ花粉を抗原としたモルモットアレルギー性鼻炎モデルを作成し，主として抗原惹起後に誘起される2相性の鼻閉ならびに抗原以外の刺激に対する鼻過敏症の発症メカニズムについて解析し，有効な治療薬の候補について検討した．遅発性の鼻閉はシステイニルロイコトリエン（CysLTs）ならびにトロンボキサン（TX）A2の受容体拮抗薬で強く抑制された．また，低濃度のLTD4およびTXA2類似薬，U-46619の点鼻は感作-惹起動物において相乗的に鼻閉を惹起することから，これらのアラキドン酸代謝物は鼻粘膜の効果器において相乗的な作用で遅発性の鼻閉を惹起している可能性が示唆された．鼻過敏性の発症はB1およびB2受容体に対する拮抗薬で抑制され，さらにこれらの受容体のアゴニストであるdes-Arg9-カリジンおよびブラジキニンの点鼻は感作-惹起動物においていずれも鼻過敏症を惹起することから，本鼻炎症状の発症にはキニン類が大きな役割を演ずる可能性が示唆された．実際の鼻アレルギー患者では，抗原惹起が繰り返され，2相性の鼻閉と鼻過敏症がオーバーラップして発症していることから，これらの鼻閉をコントロールするためには，アラキドン酸代謝物の作用を抑制することに加えて，キニン類の作用を阻害することが有効である可能性が示唆された．

Disodium cromoglycate (DSCG) is a widely used anti-asthmatic drug with few side effects. One of the mechanisms by which DSCG suppresses asthma is prevention of anaphylactic release of chemical mediators from mast cells. However, it has also been reported that DSCG attenuates late asthmatic response and airway hyperresponsiveness, the induction mechanisms of which can not be explained simply by release of chemical mediators from mast cells. Mechanisms of anti-inflammatory effects of DSCG other than its mast-cell stabilising effect have been unclear. In this study, we evaluated effects of DSCG on early and late asthmatic response in guinea pig. Guinea pigs were alternately sensitized and challenged by exposure to mists of ovalbumin+Al(OH)3 and ovalbumin, respectively. DSCG administered intratracheally before the fifth challenge dose-dependently inhibited late asthmatic response, but early asthmatic response was not affected. Interestingly, DSCG at 2 mg/kg potently suppressed increases in cysteinyl leukotrienes and thromboxane A2 in the lung during late asthmatic response. Eosinophilia was slightly reduced by DSCG. The inhibitory effect of DSCG on late asthmatic response is apparently due to inhibition of release of arachidonic acid metabolites.

Background: A toluene-2,4-diisocyanate (TDI)-induced asthma model, in which delayed-type hypersensitivity-like asthmatic airway obstruction is elicited restrictively in the lung, has never been developed. Methods: Guinea pigs were percutaneously sensitized with TDI. For the challenges, once every 2 weeks for a total of 5 times, TDI mists were delivered directly to the lung through an oral cannula, with its tip being positioned in the opening of the trachea. Time-course changes in specific airway resistance (sRaw) were measured by double-flow plethysmography. Basic mechanisms underlying TDI-induced asthma were analyzed. Results: After the 2nd–5th challenges, induction of both an early increase in sRaw that peaked at 10 min and a delayed-type sRaw elevation that peaked at 22 h were observed. Interestingly, in the sensitized/challenged animals, baseline sRaw was elevated by repeated challenge as compared to that seen for non-sensitized animals. Intratracheal administration of a bronchodilator, salbutamol, strongly suppressed the early asthmatic response (EAR) but not the delayed-type asthmatic response (DAR). During DAR, both albumin leakage and fucose secretion into the bronchoalveolar lavage fluid were increased. The cysteinyl leukotriene antagonist pranlukast failed to inhibit either EAR or DAR while the corticosteroid dexamethasone significantly suppressed DAR, without significantly affecting EAR. Conclusions: Effective delivery of TDI to the lung may induce reproducible DAR in sensitized guinea pigs with chronicity that is reflected by an increase in the sRaw baseline. DAR is not mediated by constriction of airway smooth muscles and is probably due to the concurrent presence of mucosal edema and mucus hypersecretion in the airways.

Disodium cromoglycate is an anti-asthmatic drug that has mast cell-stabilizing effects and other anti-inflammatory effects. However, the mechanisms of its anti-inflammatory effects are unclear. In this study, we evaluated effects of disodium cromoglycate on eosinophilia, early and late asthmatic responses, and production of arachidonic acid metabolites in guinea pig lungs. Guinea pigs were alternately sensitized and challenged by exposure to mists of ovalbumin+Al(OH) 3 and ovalbumin, respectively. Disodium cromoglycate (0.5–2 mg/0.1 ml/animal) administered intratracheally before the fifth challenge dose-dependently inhibited asthmatic response, but early asthmatic response was not affected. Disodium cromoglycate at 2 mg/animal potently suppressed increases in cysteinyl leukotrienes (CysLTs) and thromboxane A 2 in the lung during late asthmatic response. Eosinophilia was slightly reduced by disodium cromoglycate. The inhibitory effect of disodium cromoglycate on late asthmatic response is apparently due to inhibition of the release of arachidonic acid metabolites, some of which may be derived from eosinophils that infiltrate the lung.

The prevalence of allergic disease has increased world wide during the last decades. Pollen allergy is the most typical form of allergic disease. The increase in its frequency during recent years is the most evident. Environmental factors play an important role in the problem of pollen allergy in large cities. The aim of this research is determination of allergenicity of Canna pollen in polluted and non-polluted conditions, detection of their allergenic proteins and also elucidation of some microscopic effects of air pollutants on pollen structure and proteins. Mature and immature pollen grains of Canna indica were collected from polluted and non-polluted areas. Pollen grains were studied by scanning electron microscopy. Mice were sensitized by injection of pollen extract and an adjuvant for five times. Allergy potency of different pollen extracts were compared by means of skin test, blood eosinophills number and IgE levels in sensitized and treated animals. Pollen proteins were studied by SDS-PGE and allergenic proteins were detected by immunoblotting techniques. Scanning electron microscope study of the pollen grains showed that in polluted areas, air born particles accumulated on the surface of pollen and changed both pollen's shape and pollen's tectum. Also many vesicles were released out of polluted pollen and the pollen material agglomerated on the surface of pollen. SDS-PAGE showed that different proteins exist in mature and immature pollen. In pollen collected from polluted area, some of protein bands between 22 and 45 kDa were disappeared . Also in all polluted pollen grains, protein content of pollen decreased in response to air pollution causing the release of pollen proteins. According to our experiments and regarding induction of allergic symptoms, the polluted pollen is more effective than non-polluted one, and mature pollen has more allergy potency than immature one.

Novel 3-acetoacetylaminobenzo[b]furan derivatives having a modified triene system at the 3-position were synthesized starting with 3-aminobenzo[b]furans. The enol isomers, 3-[(3-hydroxybut-2-enonyl)amino]benzo[b]furans (), of the 3-acetoacetylaminobenzo[b]furans were obtained as stable isomers owing to formation of a hydrogen bonding between the enol hydroxyl group and the amidocarbonyl group. The planarity of the C-2 substituent through the C-3 side chain suggested the existence of a modified conjugational triene system in the enol compound. Cysteinyl leukotriene 1 and 2 receptor antagonistic activities for these compounds were evaluated. 2-(4-Cyanobenzoyl or ethoxycarbonyl)-3-[(2-cyano-3-hydroxybut-2-enonyl)amino]benzo[b]furans (, ) were moderately active.

The effects of nonsteroidal anti-inflammatory drugs (NSAIDs) on experimental allergic conjunctivitis, induced by ocular challenge with antigen in actively sensitized guinea pigs, were investigated. NSAIDs reduced the increase in prostaglandin D2 (PGD2) and E2 (PGE2) in the ocular lavage fluid. The inhibition of NSAIDs to these increases was approximately 90%-95%. NSAIDs also lowered itch-scratch response (ISR) to approximately one-third to one-half of the vehicle-treated group. However, these drugs scarcely affected plasma exudation in the conjunctiva. Ketotifen, an H1 histamine receptor antagonist, inhibited both pathophysiological changes (inhibition: 70%-80%). However, this drug was less efficacious than NSAIDs in reducing PGD2 and PGE2 levels. Moreover, topical administration of histamine induced ISR and plasma exudation; in contrast, PGD2 induced ISR exclusively. These results suggest that a part of antigen-induced ISR may be attributable to PGs. However, PGs may not play a key role in plasma exudation; other mediators such as histamine may be involved.

We evaluated roles of kinins in allergen-induced nasal blockage and sneezing, and development of nasal hyperresponsiveness to leukotriene D4 in a Japanese cedar pollen-induced allergic rhinitis model of guinea pigs. Sensitised guinea pigs were repeatedly challenged by pollen inhalation once every week. Neither a bradykinin B1 receptor antagonist, des-Arg9-[Leu8]bradykinin nor a bradykinin B2 receptor antagonist, icatibant suppressed allergen-induced sneezing and nasal blockage. However, development of nasal hyperresponsiveness to leukotriene D4 was significantly suppressed by them. The amount of bradykinin in nasal cavity lavage fluid was immediately increased after the challenge. In non-sensitised animals, hyperresponsiveness to leukotriene D4 was developed by a bradykinin B2 receptor agonist, bradykinin, but not by a bradykinin B1 receptor agonist, des-Arg10-kallidin, while in the sensitised-challenged animal, both agonists developed hyperresponsiveness. In conclusion, the nasal hyperresponsiveness appeared to be induced by kinins produced in response to the antigen challenge through activation of not only bradykinin B2 but also B1 receptors.

Studies of angiotensin-converting enzyme inhibitor-induced cough have involved extensive use of experimental models in which guinea pigs are exposed to an inhaled stimulus such as capsaicin or citric acid. In the present study, we examined enalapril-induced potentiation of spontaneous cough in guinea pigs, without an inhaled stimulus. Daily oral administration of enalapril (3 mg/kg) for 20 to 30 days enhanced spontaneous cough. This enhancement of cough was inhibited by the bradykinin B 1 receptor antagonist des-Arg 10 -[Leu 9 ]kallidin, but not by the bradykinin B 2 receptor antagonist icatibant. The amount of the bradykinin B 1 receptor agonist [ 3 H]des-Arg 10 -kallidin specifically bound to membrane fractions from the trachea and larynx was increased by prolongation of the enalapril treatment, and positively correlated well with coughing frequency. In conclusion, the present results indicate that enalapril-induced cough is mediated by up-regulation of bradykinin B 1 receptors.

To develop an allergic dermatitis model showing persistent scratching in mice, toluene-2,4-diisocyanate (TDI) was repeatedly painted onto the skin of hairless HR-1 mice, and induction of itch-associated scratching behavior was observed. When HR-1 mice were epicutaneously sensitized with 1% TDI and then challenged by repeated painting the cervicodorsal skin with 0.1% TDI once every 10 days until the 10th challenge, delayed type scratching responses peaked at 1 – 2 days after challenge. TDI at 0.1% hardly induced scratching in non-sensitized HR-1 mice. The delayed scratching response was influenced by neither an H1 nor 5-HT1/2 receptor antagonist. On the other hand, intradermal injection of histamine and serotonin induced frequent scratching in HR-1 mice. In conclusion, repeated application of TDI can induce delayed type allergic scratching. Although HR-1 mice are high responders to both histamine and serotonin, induction of the delayed response depends on neither of these chemical mediators. This delayed response may be useful in analyzing the mechanisms of allergic pruritis.

Objective and design: We evaluated the effectiveness of oral treatment with Japanese cedar pollen on experimental allergic rhinitis in guinea pigs.¶Subjects: Male Hartley guinea pigs.¶Treatment: From 16 days before the first sensitisation, 1 and 100 mg/time/animal pollen suspension was orally administered twice weekly. Animals were then sensitised and repeatedly challenged with the pollen.¶Method: Guinea pigs were sensitised by intranasal instillation of cedar pollen extracts adsorbed onto Al(OH)3 at a dose of 0.3 μg pollen protein/0.3 mg Al(OH)3/3 μl/nostril twice a day for 7 days. Then the animal was challenged by inhalation with cedar pollen (1.8 mg/nostril) once every week. We evaluated the effects of the oral treatment with antigen on: 1) sneezing frequency, 2) nasal blockage after antigen challenge, 3) nasal hyperresponsiveness to histamine and leukotriene D4, and 4) titres of anaphylactic antibodies.¶Results: During the course of the high dose administration, several animals died from a possible cytotoxicity, whereas the low dose caused no discernible change. The oral administration of the pollen at both the doses significantly inhibited nasal blockage, and the hyperresponsiveness to the stimuli was also strongly suppressed by the oral treatment. Inhibitory effectiveness did not differ substantially between the 1 and 100 mg/animal-treated groups. In contrast, neither sneezing frequency nor the increasing level of anaphylactic antibodies was influenced by the oral administration.¶Conclusions: In this study, we found that the pollen-induced nasal blockage and hyperresponsiveness were suppressed by the oral administration of the pollen in the sensitised guinea pig.

Summary Background Allergic airway eosinophilia is suppressed by cysteinyl leukotriene (CysLT) receptor (CysLT1 receptor) antagonists in several species including humans and guinea-pigs, suggesting that CysLTs are directly or indirectly involved in induction of the response. Objective We examined the effect of CysLT antagonists (pranlukast and MCI-826) on antigen inhalation-induced eosinophilia in peripheral blood and lung, and on IL-5 activity in serum during late increase of airway resistance (late asthmatic response, LAR) in sensitized guinea-pigs. Methods Guinea-pigs inhaled ovalbumin (OVA) + Al(OH)3 and OVA mists alternately for sensitization and challenge, respectively, once every 2 weeks. At the fifth challenge, the effects of CysLT antagonists and an anti-IL-5 antibody (TRFK-5) on the occurrence of LAR, and blood and lung eosinophilia, which appeared at 5 h after challenge, were examined. The time-course of IL-5 activity in the serum after the challenge was evaluated by measuring in vitro‘eosinophil survival prolongation activity’. The influence of CysLT antagonists on IL-5 activity was assessed. Results CysLT antagonists and TRFK-5 completely abolished blood and lung eosinophilia. LAR was suppressed by both MCI-826 and TRFK-5 by 40–50%. Sera obtained from sensitized, challenged animals 3 h and 4 h after challenge induced an obvious prolongation of eosinophil survival. The activity of the sera was completely neutralized by prior exposure to TRFK-5, suggesting that it reflected IL-5 activity. Increased IL-5 activity in the serum was inhibited by both pranlukast and MCI-826 by over 90%. Conclusions CysLTs produced after antigen provocation sequentially induced IL-5 production from some immune component cells via CysLT1 receptor activation. Thus, it is likely that CysLTs indirectly cause antigen-induced eosinophilia through IL-5 production.

Hairless mice fed with a special diet (named HR-AD) show atopic dermatitis (AD)-like pruritic skin inflammation that is almost completely resolved with the supplementation of an unsaturated fatty acid (UFA), the linoleic acid (LA). This suggests that the dietary deficiency of LA is the key cause of this dermatitis. However, because there is no appropriate control diet for HR-AD, the involvement of other dietary ingredients cannot be ruled out. Furthermore, it has not yet been tested whether only UFA deficiency can cause such AD-like pruritus. In this study, using semi-purified custom diets, we attempted to reproduce this syndrome. Four-week-old hairless mice were maintained on a widely used standard diet American Institute of Nutrition-76A (AIN-76A), its modifications, or HR-AD. Several modifications of fat and carbohydrate components revealed that dietary deficiency of both UFAs and cornstarch was required to induce severe skin barrier dysfunction as typically occurred in HR-AD-fed mice. An UFA- and cornstarch-deficient diet caused severe AD-like pruritus comparable to HR-AD, despite weak Th2 immune responses and absence of immunoglobulin E production. On the other hand, a diet lacking UFAs but containing cornstarch significantly alleviated the development of pruritic dermatitis. Furthermore, the supplementation of wheat starch similarly improved skin barrier function. In conclusion, this study showed that a lack of certain starches might also be the cause of diet-induced AD. Our findings could help to reproduce the diet-induced AD itch model and also provide evidence that certain starches can have protective and ameliorative effects on AD-like pruritus.

Objective and design: We investigated the effects of the thromboxane (TX) A2 antagonist seratrodast, the peptide leukotriene (p-LT) antagonist pranlukast, the antihistaminic drug terfenadine and the glucocorticoid dexamethasone on antigen-induced sneezing, biphasic nasal blockage and nasal hyperresponsiveness to histamine using a guinea pig model of allergic rhinitis.¶Subjects: Male Hartley guinea pigs were used.¶Treatment: Intranasally sensitized guinea pigs were challenged once every week for 13 weeks by inhalation of Japanese cedar pollen as the antigen. Dexamethasone and other agents were administered orally 3 and 1 h, respectively, before the 4th, 6th and 13th challenge.¶Methods: Sneezing frequency and the change in specific airway resistance (sRaw) were measured at these challenges. Two days after the 13th challenge, nasal responsiveness to histamine was evaluated by measuring sRaw after intranasal instillation of increasing doses of histamine. Moreover, the levels of TXB2, p-LTs and histamine were estimated in nasal cavity lavage fluid (NCLF) collected at the 13th challenge.¶Results: Only terfenadine (10 mg/kg) significantly inhibited sneezing at any challenge time. Seratrodast (3 and 10 mg/ kg), pranlukast (30 mg/kg) and dexamethasone (10 mg/kg), but not terfenadine, suppressed both the early and late phase elevation of sRaw (biphasic nasal blockage), although the degree of inhibition on the early phase response varied with the challenge time. In contrast, the development of nasal hyperresponsiveness to histamine was inhibited by only dexamethasone. Furthermore, biphasic increases in TXB2, p-LTs and histamine in NCLF were observed after the challenge in sensitized animals.¶Conclusions: These results suggest that TXA2 and p-LTs, but not histamine, play important roles in both the early and the late phase nasal blockage in this model of allergic rhinitis.

We examined whether nasal hyperresponsiveness to leukotriene (LT) D4 is seen in our allergic rhinitis model, which showed sneezing and biphasic nasal blockage by repeated antigen inhalation challenge, and whether a dilatation of mucosal blood vessels contributes to this hyperresponsiveness. Nasal blockage [increase of specific airway resistance (sRaw)] was indexed as nasal (hyper)responsiveness. The sensitized - challenged guinea pig showed a remarkable dose-dependent increase in sRaw by intranasal instillation of LTD4 (10 μl/nostril) at 10−10 to 10−6 M 10 h and 2 days but not 7 days after the challenge. The increase in sRaw induced by LTD4 was largely blocked by pranlukast or naphazoline, and this was dose-dependently suppressed by Nω-nitro- L -arginine methyl ester. Sodium nitroprusside induced an elevation of sRaw in the sensitized - challenged animal in the hyperresponsiveness state, but the degree did not differ from that in the non-sensitized - non-challenged group. The amount of NO2− and NO3− in nasal cavity lavage fluid after LTD4 instillation in the sensitized - challenged animal in the hyperresponsiveness state was significantly greater than that before the instillation. These results demonstrate that the hyperresponsiveness to LTD4 acquired by repeated antigen challenge is mainly due to dilatation of nasal blood vessels, which can be related to hyperproduction of nitric oxide through cysteinyl LTi-receptor activation.

We have developed an allergic rhinitis model in guinea pigs using Japanese cedar pollen as antigen. In the present study, we examined whether provocation by pollen induces similar magnitudes of rhinitis symptoms in passively and actively sensitized guinea pigs. One group of animals was actively sensitized by intranasal application of pollen extract, and another was passively sensitized by intraperitoneal injection with anti-pollen serum. Actively and passively sensitized groups were then challenged by repeated and a single pollen inhalation, respectively. In both groups, sneeze was induced immediately after the challenge. The actively sensitized animals developed not only early but also late nasal blockage, whereas the passively sensitized animals showed only early nasal blockage. In both groups, an H1 antagonist, mepyramine, inhibited the occurrence of sneezing but did not inhibit nasal blockage. Nasal hyperresponsiveness to intranasal instillation of leukotriene D4 was obvious only in the actively sensitized animals. We thus conclude that although early nasal blockage is induced by a single antigen-antibody reaction, repetitive anaphylactic reaction is required for occurrence of late nasal blockage and hyperresponsiveness to stimuli. Furthermore, histamine plays a central role in induction of sneezing but not in nasal blockage, irrespective of whether animals are actively or passively sensitized.

Objective and Design: Whether Mao-Bushi-Saishin-To (MBST), one of the formulas of classical Chinese medicine, is effective on 48-h passive cutaneous anaphylaxis (PCA) in rats and which substance in the formula is responsible for its inhibitory action were examined.¶Treatment: In the studies on PCA, MBST (hot water extract of the whole herbal formula), extracts of Ephedra herb (Mao), l-ephedrine and other reference drugs were orally administered immediately or at various times before or 5 min after the antigen challenge. In the experiments on anaphylactic histamine release from rat peritoneal mast cells, l-ephedrine and d-pseudoephedrine were added at 10-4-10-7 g/ml at 30, 10, 3 or 0 min before antigen provocation.¶Results: The time course study indicated that MBST produced a prompt and long lasting inhibition of PCA. Among the constituents of Mao, l-ephedrine exerted this prompt inhibitory activity, but d-pseudoephedrine did not. Neither pseudoephedrine nor l-ephedrine prevented the anaphylactic histamine release from isolated peritoneal mast cells.¶Conclusions: It is strongly emphasised that the rapid suppression of PCA by orally administered l-ephedrine must be exerted by a mechanism distinct from that of suppression produced following gastrointestinal absorption of the drug, because the time required for the inhibition was extraordinarily short. However, direct inhibition of anaphylactic histamine release from isolated mast cells was excluded in this inhibition of PCA.¶

We evaluated the contribution of neuropeptides to antigen-induced contractions of isolated bronchi and tracheae of passively sensitized guinea pigs using CP-96345 and SR 48968, specific antagonists of NK1 and NK2 receptors, respectively, in combination with treatment by an antihistaminic and a cysteinyl leukotriene antagonist. SR-48968 but not CP-96345, significantly inhibited the late phase of the bronchial contraction. Phosphoramidon, a neutral endopeptidase inhibitor, tended to potentiate bronchial contraction. Posttreatment with SR-48968 decreased the enhanced contraction induced by the inhibitor as well as the nonenhanced contraction to similar levels of tension. On the other hand, antigen-induced tracheal contraction was not altered by either neuropeptide antagonist. These results suggest that neuropeptides mediate the antigen-induced contractile response of the guinea pig bronchus partly through NK2 receptor stimulation.

Nasal hyperresponsiveness is one of the characteristic features of the pathogenesis of allergic rhinitis. This study examined whether repetitive inhalation of antigen (Japanese cedar pollen) led to the development of nasal hyperresponsiveness to histamine in sensitized conscious guinea-pigs. Guinea-pigs were repeatedly challenged by pollen inhalation once every week following sensitization by means of intranasal application of pollen extract plus aluminium hydroxide. The upper airways obstruction (increase in specific airway resistance (sRaw)) in response to intranasally instilled histamine was measured as an index of nasal (hyper)responsiveness. The hyperresponsiveness to histamine gradually developed with repeated pollen inhalation challenge, and the airway response at the 20th and 24th challenges was three to four orders of magnitude higher than that in nonsensitized animals. Similar degrees of hyperresponsiveness were observed at 10 h and 2 days after a pollen inhalation challenge, but the hyperresponsiveness had almost disappeared by day 7. The increased responsiveness was suppressed by pretreatment with mepyramine but not with atropine. The maximum sRaw, which was observed 10 min after histamine instillation, was largely blocked by naphazoline. Hyperresponsiveness was hardly observed on methacholine instillation. The present allergic rhinitis model, showing marked nasal hyperresponsiveness to histamine after repeated intranasal allergen challenge in guinea pigs, should be useful for investigating the pathogenesis of allergic rhinitis.

Substance P causes histamine release from rat peritoneal mast cells probably through direct activation of a specific G protein at micromolar concentrations. We found that peritoneal mast cells of a substrain of Wistar rats (Std:Wistar) responds to nanomolar concentrations of substance P by releasing histamine in a concentration-dependent manner. In addition, potent histamine release from peritoneal mast cells of the substrain rats was also induced by neurokinin A in a concentration-dependent fashion. Histamine release induced by low concentrations of substance P was significantly blocked by a tachykinin NK1 receptor antagonist, CP-96345 [(2S,3S)-cis-2-(diphenylmethyl)-N-[(2-methoxyphenyl)-methyl]-1-aza bicyclo[2.2.2]octan-3-amine dihydrochloride], whereas that induced by concentrations as high as 10 microM appeared resistant to the antagonist. The concentration-histamine release curve for neurokinin A was parallel-shifted to the right by the drug. A tachykinin NK2 receptor antagonist, SR-48968 [(S)-N-methyl-N[4-(4-acetylamino-4-phenyl piperadino)-2-(3,4-dichlorophenyl)butyl]benzamide], did not influence release stimulated by substance P and neurokinin A. On the other hand, peritoneal mast cells of Sprague-Dawley and other Wistar rats did not respond to neurokinin A. At over 1 microM but not at nanomolar concentrations, substance P caused modest histamine release from peritoneal mast cells of these rats. The results suggest that neurokinin A and nanomolar, but not micromolar concentrations of substance P stimulate tachykinin NK receptors on the peritoneal mast cells of Std:Wistar rat to release histamine.

Objective and Design: Development of nasal blockage and sneezing during repeated inhalation challenges with Japanese cedar pollens was evaluated in guinea pigs.¶Subjects: Male Hartley guinea pigs.¶Treatment: Guinea pigs were sensitized by intranasal instillation of cedar pollen extracts + Al(OH)3 2 times a day for 7 days. The animal was then forced to inhale the pollens for challenge, which was restrictively trapped in the upper airways, once a week.¶Methods: Change of specific airway resistance (sRaw), sneezing frequency, and titers of anaphylactic antibodies in the serum were measured after each of the 30 challenges.¶Results: At the first challenge, no obvious increase in sRaw was observed. However, the second and third challenges to the animals caused modest biphasic elevations of sRaw, with peaks at the first and the fourth to sixth hour. At the fourth to tenth challenges, marked elevations of sRaw were observed. However, with repetition of the inhalation challenge, the early and the late responses became almost indistinguishable because of partial overlapping as the responses expanded. All guinea pigs sneezed immediately after each pollen inhalation challenge. Apparent increases of both circulating γ1 and IgE antibodies were seen after the seventh challenge.¶Conclusions: These results indicate that the experimental allergic rhinitis established in the present study can be a valuable model for analyzing the pathogenesis of the disease and developing new therapeutic drugs.

It is well known that eosinophils increase in peripheral blood, sputum and lung tissues of patients suffering from bronchial asthma. Because eosinophil contains various cytotoxic proteins in the granule, and has abilities to produce bronchoconstrictive mediators such as peptide-leukotrienes, it has been suggested that the leukocyte involves in the occurrence of LAR as well as bronchial hyperresponsiveness, characteristic features of the disease in a serious case. In our basic experiments in vivo using a guinea pig asthmatic model, occurrence of LAR, and considerable increases of eosinophil number in the peripheral blood and the bronchoalveoli coincided with each other 5-7 hr after the antigen inhalation challenge. Significant elevation of eosinophil peroxidase activity in the lung homogenate was also observed 5 hr after the challenge. It can be considered that the increased eosinophils in the circulation, which might be recruited by cytokines such as interleukin-5 from the rapidly mobilizable bone marrow pool of eosinophils, infiltrated into the lung tissue. We also describe postulated mechanisms about eosinophil infiltration into tissues, which was recently reported, or obtained in our laboratory.

Peptide-leukotrienes (p-LTs) (LTC4, LTD4 and LTE4) are major metabolites of the arachidonate 5-lipoxygenase pathway. Although a number of papers have reported that p-LTs induce potent bronchial smooth muscle constriction as well as airway mucus secretion, vascular permeability and proliferation of airway smooth muscle cells, it had not been recognized that they are pivotal chemical mediators in asthmatic diseases until recently. Yet, several potent and selective antagonists against p-LTs and inhibitors of p-LT formation revealed that p-LTs play significant roles in not only allergic but non-allergic asthma including exercise- and aspirin-induced asthma. In addition, it has been reported that p-LTs participate in rhinitis, especially nasal blockage. In this article, recent development of drugs relating to p-LTs and their therapeutic effects for asthma are mainly reviewed.

Anaphylactic histamine release from isolated rat peritoneal mast cells was concentration-dependently blocked by a 5-min treatment with exogenous histamine at 0.9 and 9 microM and enhanced by a 20- to 30-min treatment with thioperamide (H3-antagonist) at 3 microM with significance, but little affected by mepyramine (H1-antagonist) and cimetidine (H2-antagonist) at the cell concentration of 10(6) mast cells/ml. At a low concentration of mast cells (10(4) mast cells/ml), (R)-alpha-methylhistamine (alpha-MH), an H3-agonist, at 0.9-90 microM also inhibited the release in a concentration-dependent fashion. Thioperamide, but neither mepyramine nor cimetidine, significantly restored the decreased release by alpha-MH. However, the complete restoration by thioperamide could not be achieved because the drug itself slightly but concentration-dependently inhibited anaphylactic histamine release. On the other hand, not only betahistine and dimaprit but also alpha-MH did not suppress histamine release from the mast cells induced by compound 48/80. In rat plasma, considerable levels of histamine were detected. From these results, it is strongly suggested that histamine H3-like receptors are largely responsible for the negative feedback regulation of the anaphylactic histamine release from rat peritoneal mast cells.

In this study, using a special diet-induced mouse model of atopic dermatitis, we tested the effect of chitosan-containing lotion (CL) on itch-related scratching associated with barrier-disrupted dry skin. HR-1 hairless mice fed a special diet exhibited apparent dry skin symptoms characterized by decreased skin hydration and increased transepidermal water loss. In the special diet-fed mice, scratching behavior was markedly enhanced for 60 min after oral administration of ethanol. When CL was applied once immediately after ethanol administration, the enhanced scratching response was significantly suppressed, but this effect was abolished within 30-40 min; when applied twice immediately and at 30 min, CL almost completely blocked scratching throughout 60 min. Comparison of CL and the chitosan-free vehicle showed that CL inhibited scratching more strongly and persistently than the vehicle, which transiently suppressed scratching only for 10 min after application. Although the decreased skin hydration was improved even by the vehicle, the increased transepidermal water loss was resolved only by CL. Skin surface temperature was much more reduced in CL-treated mice than in vehicle-treated mice. Collectively, CL has an antipruritic effect, which could be partly explained by recovery of skin barrier function and cooling of the skin.

Ganoderma lucidum (GL), an oriental medical mushroom, has been used in Asia for the prevention and treatment of a variety of diseases. However, the effect of GL on allergic rhinitis has not been well defined. The current study describes the inhibitory effect of GL on the biphasic nasal blockage and nasal hyperresponsiveness induced by repeated antigen challenge in a guinea pig model of allergic rhinitis. Intranasally sensitized guinea pigs were repeatedly challenged by inhalation of Japanese cedar pollen once every week. Ganoderma lucidum was orally administered once daily for 8 weeks from the time before the first challenge. The treatment with GL dose-dependently inhibited the early and late phase nasal blockage at the fifth to ninth antigen challenges. Furthermore, nasal hyperresponsiveness to intranasally applied leukotriene D₄ on 2 days after the eighth antigen challenge was also inhibited by the treatment with GL. However, Cry j 1-specific IgE antibody production was not affected by the treatment. In conclusion, we demonstrated that the pollen-induced biphasic nasal blockage and nasal hyperresponsiveness were suppressed by the daily treatment with GL in the guinea pig model of allergic rhinitis. These results suggest that GL may be a useful therapeutic drug for treating patients with allergic rhinitis.

Whether anaphylactic histamine release from rat peritoneal mast cells is influenced by betahistine, a histamine H1-receptor agonist/H3-antagonist, and dimaprit, an H2-agonist, was examined. Treatment with dimaprit at 6 and 60 microM for 20 min significantly inhibited the anaphylactic histamine release, whereas betahistine at up to 80 microM under the same conditions did not affect it. Treatment with dimaprit at 6 and 60 microM for 1 to 20 min and for 5 to 20 min, respectively, caused a time-dependent inhibition of the release, but up to 30 min treatment with 8 and 80 microM betahistine had no effect. The decreased histamine release induced by dimaprit was recovered by neither mepyramine nor cimetidine. However, thioperamide, an H3-selective antagonist, dose-dependently restored the diminished release. From these results, the inhibition of anaphylactic histamine release by dimaprit is not produced by the stimulation of H2-receptors, but involves the stimulation of H3-like receptors or H3-subtype receptors, which are distinct from the H3-receptors located in brain, and suggests that the receptor plays an important role in the negative feedback regulation of histamine release.

Although it has been suggested that prostaglandin (PG) D 2 is involved in the pathogenesis of allergic rhinitis, whether the inhibition of hematopoietic PGD 2 synthase (H-PGDS) shows beneficial effects on allergic rhinitis has been unclear. We evaluated the effects of a selective H-PGDS inhibitor, TFC-007, on nasal symptoms on Japanese cedar pollen-induced allergic rhinitis of guinea pigs. Sensitized animals were challenged with the pollen once a week. TFC-007 (30 mg/kg, p.o.) given once before a challenge almost completely suppressed PGD 2 production in the nasal tissue early and late after the challenge. Although pre-treatment did not affect the incidences of sneezing and early phase nasal blockage, late phase nasal blockage was partially but significantly attenuated; however, nasal eosinophilia was not suppressed. In contrast, when TFC-007 was given once 1.5 h after the challenge, the late phase response was not affected. Collectively, PGD 2 produced by H-PGDS early after an antigen challenge can participate in the induction of late phase nasal blockage, although the mechanism may be independent of eosinophil infilatration. The strategy for H-PGDS inhibition may be beneficial for allergic rhinitis therapy.