JP2006257040A - INHIBITOR FOR PRODUCTION OF ALLERGEN-SPECIFIC IgE ANTIBODY AND FOOD AND BEVERAGE USED FOR SUPPRESSING THE PRODUCTION OF THE ALLERGEN-SPECIFIC IgE ANTIBODY - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide an inhibitor for production of an allergen-specific IgE antibody having excellent effects for suppressing allergic symptoms of I type allergic diseases such as hay fever, all year round allergic rhinitis, atopic dermatitis, bronchial asthma and the like and having no fear of adverse effects even when ingested for a long time and food and beverages used for suppressing production of the allergen-specific IgE antibody. <P>SOLUTION: The inhibitor for the production of the allergen-specific IgE antibody contains heat sterilized mycelia of Enterococcus fecalis as an effective component and the amount of the heat sterilized mycelia of Enterococcus fecalis included in the amount of ingestion per day indicated in a product or its instruction is ≥2g. The food and beverages contain the heat sterilized mycelia of Enterococcus fecalis as the effective component. The amount of the heat sterilized mycelia of Enterococcus fecalis included in the amount of ingestion per day described in a product or its indication is ≥2g. The food and beverages are described to be used for suppressing the production of the allergen-specific IgE antibody in the product or its indication. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

The present invention relates to an allergen-specific IgE antibody production inhibitor containing heat-sterilized cells of Enterococcus faecalis as an active ingredient, and a food and drink used for inhibiting allergen-specific IgE antibody production.

  Pollen, mites, house dust, and the like are typical allergen substances, and are known to cause various allergic diseases such as hay fever, bronchial asthma, and atopic dermatitis. In particular, hay fever has many patients and has become a social problem.

  In general, for the treatment of such allergic diseases, steroidal agents, non-steroidal agents, antiallergic agents, antihistamines and the like are used externally or internally. Although these drugs have excellent antiallergic effects, their side effects are strong, so their use is limited, and it cannot be said that long-term use is preferable.

  On the other hand, in recent years, research on intestinal flora has progressed, and it has become clear that intestinal flora is closely related to the health and diseases of the host. Lactic acid bacteria and bifidobacteria are used as probiotics to prevent various diseases. There are many attempts to use it to improve symptoms.

For example, Enterococcus faecalis , which is a type of lactic acid cocci, is known to have an immunostimulatory action. An IgE antibody production inhibitor as a component is disclosed, and it is described that the daily dose of the IgE antibody production inhibitor is about 10 to 1000 mg in terms of cell weight (see paragraph 0013). .

Patent Document 2 listed below discloses histamine release in one or more bacterial cells of Enterococcus faecalis (AD101 strain) and Lactobacillus reuteri (AD0002 strain) as lactic acid bacteria isolated from human intestinal bacteria. A type I allergy inhibitor and a type IV allergy inhibitor having an inhibitory effect are disclosed. And it is described that the daily intake of the allergy inhibitor is 50 mg to 1 g as a dry weight (see paragraph 0015).
JP-A-9-2959 JP 2000-95697 A

  However, the conventional allergy suppressors containing Enterococcus faecalis as described in Patent Documents 1 and 2 as active ingredients are often free of side effects, but often do not have satisfactory effects.

  Therefore, the object of the present invention is to have an excellent inhibitory effect on the symptoms of type I allergic diseases such as hay fever, perennial allergic rhinitis, atopic dermatitis, bronchial asthma, etc. An object of the present invention is to provide an allergen-specific IgE antibody production inhibitor that is free from worries of side effects, and foods and drinks that are used to suppress allergen-specific IgE antibody production.

In order to achieve the above object, the allergen-specific IgE antibody production inhibitor of the present invention contains heat-killed cells of Enterococcus faecalis as an active ingredient, and per day indicated on the product or its instructions. heat sterilization cell amount of the Enterococcus faecalis contained in intake (Enterococcus faecalis) is characterized in that there is a more 2g of.

In the allergen-specific IgE antibody production inhibitor, the amount of heat-sterilized cells of Enterococcus faecalis contained in the daily intake is preferably 2 to 10 g.

The Enterococcus faecalis is preferably FERM BP-10284 (AIST / patent microorganism deposit center deposit number).

  The allergen-specific IgE antibody production inhibitor is preferably one that suppresses eosinophil infiltration, and further suppresses IL-4 production while enhancing interferon-γ production. preferable.

On the other hand, the food / beverage product of the present invention contains Enterococcus faecalis heat sterilized bacteria as an active ingredient, and the Enterococcus faecalis contained in the daily intake indicated on the product or its instructions. ( Enterococcus faecalis ) has a heat-sterilized bacterial mass of 2 g or more, and is characterized in that a label indicating that it is used for suppression of allergen-specific IgE antibody production is attached to the product or its instructions.

In the food and drink, it is preferable that the amount of heat-sterilized cells of Enterococcus faecalis contained in the daily intake is 2 to 10 g.

The Enterococcus faecalis is preferably FERM BP-10284 (AIST / patent microorganism deposit center deposit number).

  In addition, it is preferable that the product or its instructions are further labeled with an indication that they are used to suppress eosinophil infiltration, and further, while inhibiting IL-4 production, interferon-γ It is preferable that a label indicating that it is used for enhancing production is attached.

  The allergen-specific IgE antibody production inhibitor and food and drink of the present invention have an amount of heat-sterilized Enterococcus faecalis contained in the daily intake indicated on the product or its instructions is 2 g or more, Makes it possible to reliably ingest heat-sterilized cells of Enterococcus faecalis that can be sufficiently expected to have a physiologically active effect. Type I allergies such as hay fever, perennial allergic rhinitis, atopic dermatitis, bronchial asthma, etc. It is expected to have an excellent inhibitory effect on symptoms of sexually transmitted diseases.

  According to the present invention, by suppressing the production of allergen-specific IgE antibodies, excellent suppression of symptoms of type I allergic diseases such as hay fever, perennial allergic rhinitis, atopic dermatitis, bronchial asthma, etc. An allergen-specific IgE antibody production inhibitor and food and drink that can be expected to have an effect can be provided. Since the allergen-specific IgE antibody production inhibitor and food and drink of the present invention are highly safe, they can be suitably used for children and children.

  In addition, by suppressing the infiltration of eosinophils, it can be expected to suppress the allergic reaction by suppressing the release of chemical mediators.In addition, it suppresses IL-4 production while enhancing interferon-γ production, thereby increasing Th1 / Th2 balance improvement can be expected.

Enterococcus faecalis used in the allergen-specific IgE antibody production inhibitor and foods and drinks of the present invention includes FERM BP-10284 (National Institute of Advanced Industrial Science and Technology, Patent Microorganism Depositary Depositary Number, display for identification) Enterococcus faecalis EC-12 ), ATCC 19433, ATCC 14508, ATCC 23655, IFO 16803, IFO 16804 and the like can be exemplified, and among these, the FERM BP-10284 strain can be preferably exemplified.

In addition, enterococcus faecalis heat-killed cells can be recovered from a culture obtained by culturing Enterococcus faecalis according to a conventional method, for example, by filtration or centrifugation. After washing with water, it can be prepared by suspending in water or the like, heat-treating at 80 to 120 ° C. for 3 seconds to 30 minutes, and then concentrating and drying as necessary. Since the sterilized bacterial powder by heat treatment of Enterococcus faecalis ( FERM BP-10284 ) is commercially available under the trade name “EC-12” (manufactured by Combi Co., Ltd.), such a commercial product can also be used.

In the allergen-specific IgE antibody production inhibitor of the present invention, the amount of heat-sterilized cells of Enterococcus faecalis contained in the daily intake shown in the product or its instructions is 2 g or more, Preferably it is 2-10g, More preferably, it is 4-8g.

If the amount of Enterococcus faecalis contained in the daily intake is less than 2 g, a sufficient antiallergic effect cannot be expected, and if it exceeds 10 g, a tender stool tendency may be exhibited. The number of cells per 2 g of heat-sterilized cells of Enterococcus faecalis (FERM BP-10284) is 1 × 10 ¹³ .

  Further, in the present invention, “displayed on the product” means a product directly displayed on a packaging material such as a container, bag, box, etc. of the product, and “displayed in its instructions” It means a printed matter enclosed in a product bag or box, or a product brochure.

  The allergen-specific IgE antibody production inhibitor of the present invention is supplemented with excipients, sweeteners, acidulants, vitamins, minerals, oligosaccharides, dietary fiber, etc., as necessary, in addition to the above-mentioned cells. And can be prepared as a powder, granule, tablet, capsule, liquid, paste or jelly.

In addition, the food and drink used for suppressing the production of allergen-specific IgE antibody of the present invention contains the heat sterilized cells of Enterococcus faecalis as an active ingredient, and is displayed on the product or its instructions. heat sterilization cell amount of the Enterococcus faecalis contained in daily intake (Enterococcus faecalis) are equal to or greater than 2g, product or that used for allergen-specific IgE antibody production suppressing its manual Is attached.

  The type of food or drink of the present invention is not particularly limited, but is a powder, granule, tablet, capsule, health food or drink prepared in a liquid, paste or jelly form, various noodles such as bread, udon and ramen, and natto Examples include soft drinks, various drinks such as drinks and jelly drinks, various instant foods such as jelly, gummi, candy, biscuits, instant soup and instant miso soup, retort foods, foods for microwave ovens, and the like.

In the food and drink of the present invention, the amount of heat-sterilized cells of Enterococcus faecalis contained in the daily intake is 2 g or more, preferably 2 to 10 g, more preferably 4 ˜8 g.

There is no restriction | limiting in particular in the addition method of the heat-sterilized microbial cell of Enterococcus faecalis in the said food / beverage products, It can add from the beginning together with the other raw material used for each food / beverage product. Therefore, even if it adds to the food-drinks which require heat processing (process under temperature conditions that kill bacteria, such as heat sterilization, baking, and steaming), an effect is not impaired.

  Using commercially available Enterococcus faecalis heat sterilized bacterial powder (trade name “EC-12”, manufactured by Combi Co., Ltd., hereinafter abbreviated as “EC-12”), the antiallergic effect on asthma-induced model mice is as follows: Investigated by the method.

(1) Animals Seven-week-old female BALB / c mice (purchased from Japan SLC) were divided into 3 groups (6 per group).

(2) Preparation of sample “EC-12” is suspended in distilled water to prepare sample 1 (concentration 0.4 mg / ml), and 0.5 ml is orally administered daily (200 μg / mouse) to test group 1 during the test period. did. Sample 2 (concentration 4 mg / ml) was prepared in the same manner, and 0.5 ml was orally administered daily (2 mg / mouse) to Test Group 2. In addition, 0.5 ml of sterile water was orally administered daily to the control group.

(3) Asthma-Inducing Model Mice A mixture of 100 μg of ovalbumin (OVA) and 1.6 mg of aluminum hydroxide gel was subcutaneously administered on the 0th and 7th days to each group of mice. On days 14, 15 and 16, OVA 10 μg was administered nasally. Dissected on day 18 and serum, lung lavage fluid, lung and spleen were collected.

(4) Preparation and culture of splenocytes Spleen cells were adjusted to 2x10 ⁶ / ml, stimulated for 3 days with anti-CD3 monoclonal antibody (mAb) (145-2C11) (Nippon Becton Dickison), and cultured. The amount of IFN-γ and IL-4 in the serum was examined by ELISA.

(5) Measurement of antibody titer and cytokine Antibody titer and IFN-γ were serially diluted in a 96-well plate coated with OVA or anti-mouse IFN-γ mAb (R4-6A2) (Nippon Becton Dickison) Was added. As detection antibodies, AP-labeled anti-mouse IgG1 (manufactured by ICN), IgG2a antibody (manufactured by ZYMED) or biotin-labeled anti-mouse IFN-γ antibody (AN-18) (manufactured by Nippon Becton Dickyson) and AP-labeled streptavidin ( Using ZYMED), color was developed using p- nitrophenyl phosphate as a substrate, the reaction was stopped with 3N NaOH, and the absorbance at 450/540 nm was measured with a microplate reader.

IgE was coated with anti-mouse IgE mAb (Nippon Becton Dickinson) and serially diluted samples were added. For detection, biotin-labeled OVA and HRP-labeled streptavidin (manufactured by ZYMED) were used to develop color under the trade name “TMB Microwell Peroxidase” (manufactured by Funakoshi Co., Ltd.), and the reaction was stopped with 2N H ₂ SO ₄ , followed by absorbance at 450/540 nm. Was measured.

IL-4 was measured using the trade name “OptEIA ^™ Set” (BD Bioscience).

(6) Measurement of cell number The total leukocyte count and eosinophil count in the lung lavage fluid on the 18th day of the asthma induction model mouse were measured. The total white blood cell count was stained with Turku's solution and measured with a hemocytometer, and eosinophils were stained with Giemsa stain and measured with a microscope.

(7) Lung tissue section Lungs were collected from each mouse and fixed with 4% paraformaldehyde. After embedding in paraffin, the slices were stained with Hematoxylin-Eosin.

(8) Results FIG. 1 shows the results of measuring the total white blood cell count and eosinophil count in the alveolar lavage fluid. FIG. 1 shows that the number of eosinophils is decreased in test group 1 and test group 2 as compared to the control group. In particular, test group 2 showed a significant difference (p <0.001) from the control group. On the other hand, there was no difference in the total leukocyte count in each group.

  As a result of measuring the amount of IFN-γ in the culture supernatant after in vitro stimulation of splenocytes on day 18 of an asthma-induced model mouse with anti-CD3 mAb for 3 days, the amount of IL-4 was measured. The results are shown in FIG. 2 and 3, it can be seen that in test group 2, the amount of IFN-γ tends to increase and the amount of IL-4 tends to decrease compared to the control group. On the other hand, in the test group 1, a remarkable tendency was not seen.

  The results of measuring the serum OVA-specific antibody titer on day 18 of asthma-induced model mice are shown in FIG. FIG. 4 shows that in the test group 2, the amounts of IgG1, IgG2a, and IgE are significantly reduced, and in particular, the amount of IgE is significantly reduced. On the other hand, in Test Group 1, it can be seen that the amounts of IgG2a and IgE tend to decrease.

  The results of pathological comparison of the lungs of the mice of the control group and test group 2 are shown in FIG. From FIG. 5 (A), leukocyte infiltration in the inflamed area was observed in the control group. On the other hand, from FIG. 5 (B), in the test group 2, infiltration of leukocytes was remarkably suppressed.

  From the above results, asthma model mice were orally administered with a predetermined amount (2 mg / mouse) of heat sterilized cells of Enterococcus faecalis 1) eosinophil infiltration was significantly suppressed, and 2) Th2 cytokines were It was found that the antibody response involved was also remarkably suppressed, and 3) the amount of IFN-γ produced by splenocytes increased and IL-4 production was suppressed.

  Here, Th1 / Th2 balance is one of the important factors for maintaining homeostasis, and it is known that asthmatic patients have a T2-dominant state of T cell responses. However, it was suggested that oral administration of a predetermined amount of Enterococcus faecalis heat-sterilized bacterial powder restores Th1 / Th2 balance through macrophage activation and improves the disease state.

  The allergen-specific IgE antibody production inhibitor and food and drink of the present invention can be used not only as a health food but also as a functional food material that can be easily added to various foods and drinks. Moreover, since there is no worry of side effects, it can be suitably used particularly for children and children.

It is a figure which shows the result of having measured the total white blood cell count and the number of eosinophils in the alveolar lavage fluid of each group of mice. It is a figure which shows the result of having measured the amount of IFN-gamma in the culture supernatant after stimulating the spleen cell of each group mouse | mouth with anti-CD3 mAb for 3 days in vitro. It is a figure which shows the result of having measured the amount of IL-4 in the culture supernatant after spleen cells of each group of mice were stimulated with anti-CD3 mAb for 3 days in vitro. It is a figure which shows the result of having measured the OVA specific antibody titer in the serum of the mouse | mouth of each group. It is a figure which shows the tissue slice of the lung of the mouse | mouth of a control group and the test group 2. FIG.

Claims (10)

Containing heat killed cells of Enterococcus faecalis (Enterococcus faecalis) as an active ingredient, heat sterilization cells of the contained in the product or intake per day which is displayed on the manual Enterococcus faecalis (Enterococcus faecalis) An allergen-specific IgE antibody production inhibitor characterized by having an amount of 2 g or more. The allergen-specific IgE antibody production inhibitor according to claim 1, wherein the amount of heat-killed cells of Enterococcus faecalis contained in the daily intake is 2 to 10 g. The Enterococcus faecalis (Enterococcus faecalis) is FERM BP-10284 (AIST-Patent Organism Depositary accession number) allergen specific IgE antibody production inhibitor according to claim 1 or 2 wherein the.   The allergen-specific IgE antibody production inhibitor according to any one of claims 1 to 3, which suppresses eosinophil infiltration.   The allergen-specific IgE antibody production inhibitor according to any one of claims 1 to 4, which suppresses IL-4 production and enhances interferon-γ production. Containing heat killed cells of Enterococcus faecalis (Enterococcus faecalis) as an active ingredient, heat sterilization cells of the contained in the product or intake per day which is displayed on the manual Enterococcus faecalis (Enterococcus faecalis) A food or drink characterized in that the amount is 2 g or more, and a label indicating that the product or its instruction is used for suppressing the production of allergen-specific IgE antibody. The food or drink according to claim 4, wherein the amount of heat-sterilized cells of Enterococcus faecalis contained in the daily intake is 2 to 10 g. The Enterococcus faecalis (Enterococcus faecalis) the claims 6 or 7, wherein the food or beverage is FERM BP-10284 (AIST-Patent Organism Depositary accession number).   The food or drink according to any one of claims 6 to 8, wherein a label indicating that the product or its instruction is further used to suppress eosinophil infiltration is attached.   The allergen-specificity according to any one of claims 6 to 10, wherein the product or instructions thereof further indicate that the product is used to enhance IL-4 production while enhancing interferon-γ production. Food and drink used to suppress the production of target IgE antibodies.