Inhibitory effects of apple polyphenol extract (AP) and procyanidin contained in AP on in vitro pancreatic lipase activity and in vivo triglyceride absorption in mice and humans were examined. AP and procyanidin considerably inhibited in vitro pancreatic lipase activity. However, polyphenols, except for procyanidin, in AP (i.e., catechins, chalcones, and phenol carboxylic acids) showed weak inhibitory activities on pancreatic lipase. Procyanidins separated by normal-phase chromatography according to the degree of polymerization were also examined. Inhibitory effects of procyanidins increased according to the degree of polymerization from dimer to pentamer. On the other hand, pentamer or greater procyanidins showed maximal inhibitory effects on pancreatic lipase. These results suggested that with respect to in vitro pancreatic lipase inhibition, the degree of polymerization was an important factor and oligomeric procyanidin mainly contributed. Next, we performed a triglyceride tolerance test in mice and humans. Simultaneous ingestion of AP and triglyceride significantly inhibited an increase of plasma triglyceride levels in both models. These results suggested that the oligomeric procyanidins contained in AP inhibited triglyceride absorption by inhibiting pancreatic lipase activity in mice and humans.

A major problem with allergen-specific immunotherapy involving repeated injection of allergens is the risk of an anaphylactic reaction. We engineered the major house dust mite allergen, Der f 2, to reduce its capacity to induce skin test reactivity and histamine release from peripheral blood basophils in allergic patients. The engineered allergen, in which the disulfide bond that linked the N- and C-terminal sequences of Der f 2 was disrupted, retained T-cell epitopes essential for immunotherapy and ability to stimulate T-cell proliferation. Such engineered allergens are potentially useful for safer and more effective immunotherapy for allergies.

Background: Monkeys are considered to have an immune system very similar to that of humans, as compared with mice, rats, and guinea pigs. Although primate allergic models to several pollen allergens have been developed, no model of house dust mite allergy has been reported. In this study, we attempted to induce type I allergy to mite allergens in rhesus monkeys. Methods: Six rhesus monkeys were immunized subcutaneously with crude mite extract adsorbed on aluminum hydroxide for 4 months. Then 5 monkeys positive for IgE production to mite extract were further immunized subcutaneously and conjunctivally with recombinant Der f 2 (rDer f 2). The status of sensitization to mite extract and rDer f 2 in monkeys was examined before and after the immunization. Plasma antigen-specific IgE and IgG levels, cutaneous reaction, and histamine release from peripheral blood leukocytes were measured. After conjunctival immunization, immediate conjunctivitis and leukocyte influx into conjunctiva after rDer f 2 challenge were examined. Results: After immunization with crude mite extract, 5 of 6 sensitized monkeys showed IgE response to the mite, and 4 out of 5 rDer f 2-sensitized monkeys exhibited IgE production to rDer f 2. Three monkeys sensitized with rDer f 2 showed immediate conjunctivitis and conjunctival eosinophilia after applying rDer f 2 to their eyes. Sensitized animals also showed IgG response to mite antigens. Conclusion: Four rhesus monkeys were positive for IgE production and allergic reactions to both mite extract and rDer f 2. These monkeys could represent a useful model for studying the development and regulation of house dust mite allergy.

SUMMARYC8/119S is a mutant of recombinant Der f 2 (rDer f 2), and lacks a disulphide bond possessed by wildtype rDer f 2. In humans and mice, C8/119S has a very weak IgE-binding capacity compared with the wild-type, but possesses a T cell reactivity comparable to that of the wild-type. C8/119S may thus be a safe immunotherapeutic agent for house dust mite allergy. The aim of the present study was to evaluate whether the intranasal administration of C8/119S could suppress an immediate allergic reaction in mice sensitized with wild-type rDer f 2, possessing an allergic activity comparable to native counterparts purified from mite extract. Seven-week-old male A/J mice were immunized with wild-type rDer f 2 four times, and then intranasally administered 0·2-2 mg of wild-type, 0·2-20 mg of C8/119S, or PBS alone, three times a week for 4 weeks. Seven days after the last administration, the mice were examined for an immediate allergic reaction. The animals administered 2 mg of C8/119S (C2.0 group) showed significantly reduced immediate bronchoconstriction provoked by the i.v. injection of 1 and 10 mg of wild-type rDer f 2, compared with the PBS-treated mice. Similar results were obtained when we examined mice 10 weeks after the last administration. The reactions in the other groups given wild-type or C8/119S also tended to decrease in severity in comparison with the animals of the PBS group. The allergic phenotypes of the T cells, B cells, and basophils in the C2.0 group were shifted to that of naive mice without immunization. We conclude that C8/119S has hyposensitizing activities in mice sensitized with wild-type rDer f 2. C8/119S may be useful for immunotherapy of house dust mite allergy.

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We investigated the effects of Cimicifuga racemosa (CR) plant extracts on the changes in levels of the cerebral monoamines norepinephrine (NE), dopamine (DA), and serotonin (5-HT), the respective metabolites 3-methoxy-4-hydroxyphenylglycol (MHPG), 3,4-dihydroxyphenylacetic acid (DOPAC), and 5-hydroxyindoleacetic acid (5-HIAA), and plasma corticosterone in mice subjected to acute immobilization stress. Single oral administration of the CR extract (1,000 mg/kg) significantly attenuated plasma corticosterone levels that had been increased as a result of enforced immobilization. Acute immobilization stress caused significant changes in the corresponding amine-to-metabolite ratios in the hypothalamus, hippocampus, and cortex; however, CR-extract treatment significantly attenuated the MHPG/NE change in the hypothalamus, and the 5-HIAA/5-HT changes in each region of the brain. Our results suggest that the CR extract interacts not only with the hypothalamic-pituitary-adrenal (HPA) axis but also with the sympathetic adrenomedullary (SAM) system under stress conditions. Thus the CR extract can alleviate acute stress responses by suppressing the changes of amine-to-metabolite ratio in brain.Cimicifuga racemosa (CR) is a member of the Ranunculaceae family that is indigenous to eastern North America, with a range that extends as far as south Florida. Dried CR rhizomes are known as black cohosh and have been widely used as herbal dietary supplements for almost five decades. Historically, Native American women ingested CR water extract for pain relief during menstruation and childbirth. In recent years, ethanolic and isopropanolic extracts of CR have been used for the treatment of general menopause symptoms including hot flushes, profuse sweating, irritability, and anxiety, and its popularity has grown among women hoping to avoid the potential toxicity of classical hormonereplacement therapy (2). We previously showed that CR extracts had antistress activities in mice in which plasma corticosterone and aspartate aminotransferase (AST) levels were increased as a result of enforced immobilization: a single oral administration of CR extract (1,000 mg/kg) significantly attenuated these increases (23). Previous studies have reported that acute immobilization stress not only elicits activation of the hypothalamic-pituitary-adrenal (HPA) axis but also affects the metabolism of monoamines such as norepinephrine (NE), dopamine (DA), and serotonin (5-HT) in the brain. And these amine-to-metabolite ratios are increased by acute immobilization stress. Furthermore, the relationships among NE, DA, and 5-HT play an important role in regulation of the sympathetic adrenomedullary (SAM) system under stress conditions (5,20,31). Several studies have reported that the neuronal activities of serotonergic and dopaminergic pathways innervating the striatum, hippocampus, hypo-

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Recombinant Der f2 (rDer f2) has recently been developed as a promising allergen for the diagnosis and immunotherapy of house-dust mite allergy, and studies in immunology. The aim of the present study was to evaluate whether oral administration of rDer f2 could suppress an immediate allergic reaction in mice sensitized with mite allergen. We developed a murine allergic model that showed bronchoconstriction after inhalation of rDer f2, and studied the effect of oral administration of rDer f2 on the reaction.Seven week old male A/J mice were intranasally immunized with rDer f2 12 times. Sensitized mice showed anti-rDer f2 immunoglobulin (Ig)E production and immediate airway constriction after inhalation of 10 mg·mL -1 of rDer f2, as determined by the Konzett-Rössler method. Immunized animals were divided into three groups, and fed phosphate-buffered saline (PBS), 0.1 mg·day -1 , or 1 mg·day -1 of rDer f2 for 4 weeks, respectively. Seven days after the last feeding, the mice were examined for their immediate response.Animals fed with 1 mg·day -1 rDer f2 showed significantly reduced bronchoconstriction after inhalation of both 2 mg·mL -1 and 10 mg·mL -1 of rDer f2 compared with PBS-fed mice. Similar results were obtained when we examined mice 10 weeks after the last feeding. Reactions in the 0.1 mg·day -1 rDer f2-fed group also tended to decrease in comparison with PBS-fed animals. Plasma anti-rDer f2 IgE, IgG1, IgG2a, and IgG2b levels were not changed by feeding with rDer f2.We conclude that recombinant Der f2 exhibits both sensitizing and hyposensitizing activities in mice. rDer f2 may be useful in immunotherapy and diagnosis of housedust mite allergy. Eur Respir J 1998; 11: 144-150 To date, several clinically important allergens such as Der 1 and Der 2 have been identified in crude mite extract [1][2][3][4]. Among others, the structural gene of Der f2, one of the major mite allergens found in Dermatophagoides farinae was cloned, and the product (recombinant Der f2 (rDer f2)) shown to exhibit allergic activity in humans comparable with native Der f2 prepared from crude mite extract [5,6]. Since Der f2 has been shown to sensitize over 80% of patients with house-dust mite allergy, rDer f2 should prove useful in the diagnosis and immunotherapy of allergic asthma and rhinitis caused by house-dust mites. It should also provide a basis for research into allergies and immunology.In the present study, with special reference to the therapeutic application of rDer f2, we established a murine allergic model by intranasal or intraperitoneal injection of rDer f2 to A/J mice, which are highly responsive to Der f2 [7]. We examined the effect of oral-hyposensitization therapy with rDer f2 against the immediate bronchoconstriction in the intranasally sensitized animals because their reaction after rDer f2 inhalation was more intense than that of intraperitoneally immunized mice. Materials and methodsAntigen rDer f2, clone 1 [5], was kindly supplied by the Nikka Whisky Distilling Co., Ltd. (Chiba, Japan), and dissolved in...