Lactobacillus plantarum strain, culture solution derived from same and anti-allergic application of lactobacillus plantarum strain

Abstract

Relates to a novel microorganism, a lysate thereof, a culture solution, a culture solution extract and an anti-allergic application thereof. The novel strain according to one aspect and a lysate, a culture solution, a culture solution extract, or a mixture thereof derived therefrom have an effective effect for preventing, ameliorating, or treating allergic diseases.

Description

Technical Field

[0001] The invention relates to a novel microorganism, its lysate, culture fluid, culture fluid extract and anti-allergic use thereof. Background Art

[0002] Microbiome refers to the microorganisms that exist in a specific environment and their entire genetic information. It refers to the collection of genomes that represent the entire genetic information of a single organism. Therefore, the human microbiome refers to the microorganisms that live inside and outside the human body and their entire genetic information.

[0003] The human body is in a symbiotic relationship with many microorganisms. In particular, the intestine is the best environment for microorganisms to absorb nutrients and form systemic colonies, so the largest number of microorganisms are present there. Intestinal microorganisms provide nutrients that the host cannot produce through its own enzymes alone, and are closely related to the host's metabolism and immune system. It has been reported that it is related to the occurrence of various diseases such as irritable bowel syndrome, obesity, atopy, depression, rheumatoid arthritis, autism spectrum disorder, and dementia.

[0004] Allergy includes not only IgE-mediated allergic hypersensitivity reactions but also non-IgE-mediated hypersensitivity reactions. IgE-mediated allergic diseases are allergic reactions to food caused by IgE allergic antibodies, and non-IgE-mediated allergic diseases are diseases that occur due to reactions involving other components of the immune system in addition to IgE antibodies. The mechanism of non-IgE-mediated allergic diseases is unclear, but it is known that innate immunity and cellular immunity (T lymphocyte processes) are involved, but IgE antibodies are not actually involved. It is known that the activation of mast cells induces mast cell degranulation, so that chemical mediators such as histamine, cytokines and chemokines present in the granules are secreted outside the cells, which induces increased vascular permeability, smooth muscle contraction and increased mucus secretion, causing type I allergic reactions.

[0005] The mast cells produce various cytokines through the high-affinity IgE receptor (FcεRI), thereby promoting the expression of inducers of inflammatory reactions and inducing inflammatory allergic reactions. Among them, IL-4 is generated in T lymphocytes or mast cells, etc., induces the generation of IgE that induces type I allergic reactions, and regulates the mediated immune response. It has been reported that the generation of IL-4 increases in actual patients (Ngoc et al., 2005). In addition, when an allergic reaction begins in the body, TNF-α degranulates from mast cells, flows out together with histamine and other inflammatory mediator factors, and induces the generation of other cytokines and inflammatory reaction mediator factors, leading to further deterioration of allergies and inflammatory reactions (Chung and Barnes, 1999). The histamine exists before an allergic reaction and is released to the outside when stimulated by an allergy inducer (Schwartz and Huff, 1998).

[0006] Allergy is a disease in which substances that have no effect on ordinary people induce hypersensitivity reactions such as urticaria, itching, runny nose, cough, etc. only in certain specific people due to the dysfunction of the immune system. It can be induced by environmental factors such as environmental pollution, westernization of diet and lifestyle, and stress, as well as genetics and various allergens. The prevalence rate globally has a tendency to increase, and 20% to 25% of the national population suffers from allergic diseases such as rhinitis, asthma, atopic dermatitis, and food allergies, etc. Recently, there is evidence indicating that it is also related to late-phase reactions and chronic allergic reactions, and thus it is known to be involved in overall allergic inflammation.

[0007] To treat allergic diseases, various methods for treating allergic diseases have been proposed, such as avoiding allergens, administering anti-allergy drugs, regulating the synthesis of IgE in the body, developing anti-IgE antibodies, etc. However, the above methods also have many drawbacks, such as being unable to fundamentally treat the cause of allergies, insufficient drug efficacy, or occurrence of serious side effects, etc.

[0008] Therefore, there is an urgent need to discover anti-allergy active substances for developing allergy therapeutics. To demonstrate anti-allergy activity, it is not enough to merely have IgE-mediated allergy treatment activity, and it must also have treatment activity against non-IgE-mediated allergies.

[0009] Recently, as a result of conducting a metagenomic project to identify the intestinal bacterial flora, the correlation between the distribution of the intestinal flora and various immune system diseases including allergies has been revealed, and currently, it is in the stage of identifying the distribution and diversity of intestinal bacteria and the close relationship between intestinal immunity and the immune system. It is necessary to develop a substance that has treatment activity against IgE-mediated allergies and treatment activity against non-IgE-mediated allergies by utilizing the results of identifying the intestinal bacterial flora. Summary of the Invention Technical Problem

[0010] On the one hand, a composition is provided which inhibits the production of cytokines mediated by helper T cells type 2 (Th2) or alleviates allergic reactions, and the composition includes a Lactiplantibacillus plantarum strain belonging to the genus Lactiplantibacillus as an active ingredient.

[0011] On the one hand, a Lactiplantibacillus plantarum GBCC_F0070 strain belonging to the genus Lactiplantibacillus is provided, and its deposit number is KCTC15051BP.

[0012] On the other hand, a lysate, a culture solution or an extract of the culture solution derived from the strain is provided.

[0013] On the other hand, a health food for preventing or improving allergic diseases is provided, which includes a Lactiplantibacillus strain, a lysate, a culture solution or a mixture thereof derived from the strain as an active ingredient.

[0014] On the other hand, a pharmaceutical composition for treating or preventing allergic diseases is provided, which includes a Lactiplantibacillus strain, a lysate, a culture solution or a mixture thereof derived from the strain as an active ingredient.

[0015] On the other hand, a cosmetic composition for preventing or improving skin allergic diseases is provided, which includes a Lactiplantibacillus strain, a lysate, a culture solution or a mixture thereof derived from the strain as an active ingredient.

[0016] On the other hand, a method for preventing or treating allergic diseases is provided, which includes the step of administering an effective amount of a composition to an individual in need thereof, and the composition includes the Lactiplantibacillus plantarum GBCC_F0070 strain with the deposit number of KCTC15051BP, a culture of the strain, a lysate of the strain or a mixture thereof.

[0017] On the other hand, the use of a composition including the Lactiplantibacillus plantarum GBCC_F0070 strain with the deposit number of KCTC15051BP, a culture of the strain, a lysate of the strain or a mixture thereof in the manufacture of a pharmaceutical preparation or a health food preparation for preventing or treating allergic diseases is provided. Technical Solution

[0018] On the one hand, a composition is provided that inhibits the production of cytokines mediated by helper T cells type 2 (Th2) or alleviates allergic reactions, and the composition includes a Lactiplantibacillus plantarum strain belonging to the genus Lactiplantibacillus as an active ingredient.

[0019] On the one hand, a Lactiplantibacillus plantarum strain belonging to the genus Lactiplantibacilus is provided.

[0020] Lactiplantibacillus is a microorganism belonging to the genus of aerobic or facultative anaerobic Gram-positive bacilli widely distributed in nature. Microorganisms belonging to the genus Lactiplantibacillus include Lactiplantibacillus plantarum and fermentum, etc. As a result of research on developing new strains with excellent anti-allergic effects, the present inventors selected Lactiplantibacillus plantarum GBCC_F0070 as a candidate anti-allergic strain. The strain was deposited at the Korea Research Institute of Bioscience and Biotechnology Biological Resource Center on August 16, 2022, under the accession number KCTC15051BP. The strain corresponds to a probiotic strain, is harmless to the human body, and is used without side effects.

[0021] In a specific embodiment, the Lactiplantibacillus plantarum strain may be the strain deposited under the accession number KCTC15051BP.

[0022] In a specific embodiment, the Lactiplantibacillus plantarum strain may be a strain including the 16S rRNA of SEQ ID NO: 1.

[0023] In a specific embodiment, the strain may be a strain having a 16S rRNA composed of the nucleotide sequence of SEQ ID NO: 1 or including a 16S rRNA having a nucleotide sequence homology of 97% or more therewith. Specifically, it has at least 93%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9% or 100% homology with the nucleotide sequence composed of SEQ ID NO: 1 in this specification.

[0024] In a specific embodiment, the Lactiplantibacillus plantarum strain may be a strain including the recA gene sequence of SEQ ID NO: 2.

[0025] In a specific embodiment, the strain may be a strain having the recA gene sequence of SEQ ID NO: 2 or a recA including a gene sequence having 97% or more gene sequence homology therewith. Specifically, it has at least 93%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9% or 100% homology with the gene sequence consisting of SEQ ID NO: 2 in this specification.

[0026] In a specific embodiment, the strain may be a strain that is a mutant of a naturally occurring strain.

[0027] In a specific embodiment, the strain may be a live bacterium, a dead bacterium, an attenuated bacterium, or a cytoplasmic fraction obtained by disrupting the strain, and preferably may be a live bacterium.

[0028] In a specific embodiment, the strain may have anti-allergic activity.

[0029] Specifically, without being limited to a particular theory, the allergy induction mechanism can be divided into a first-stage mechanism and a second-stage mechanism.

[0030] In the first stage, an external allergen first penetrates into the body through epithelial cells. The penetrated allergen is phagocytosed by antigen-presenting cells, resulting in a part of its fragments being presented to T cells. At this time, the T cells receiving the antigen differentiate into type 2 helper T cells (Th2), so that Th2-mediated cytokines, such as interleukin (IL)-4, etc., can be generated. The generated cytokines can activate B cells to induce the generation of immunoglobulin E (IgE) from B cells. At this time, the generated IgE antibody binds to the FcεRI receptor present on the surface of mast cells, and this state is called the atopic state, and it can be said to be a preparatory (waiting) state for allergy induction.

[0031] The second stage may be as follows: The allergen that penetrated and was remembered by immune cells in the first stage re-penetrates and cross-binds with IgE bound to mast cells, thereby inducing the degranulation process of mast cells, and thus releasing allergic inflammatory factors such as histamine, leukotrien, and tryptase present in mast cells to the outside. These substances usually include histamine, which may be the direct cause of various allergic inflammatory diseases. As described above, the mechanism of action of current therapeutic agents or allergy improvement materials may focus on blocking the release of histamine from mast cells, which is the final stage of allergy, and the action of histamine.

[0032] In a specific embodiment, the strain may have the activity of inhibiting Th2-mediated cytokine production, inhibiting the degranulation activity of mast cells, inhibiting the Ig level in serum, or inhibiting the MCPT-1 level in serum. Specifically, the inhibiting mast cell degranulation activity means that it can inhibit the secretion of β-hexosaminidase from activated mast cells.

[0033] In a specific embodiment, the strain may reduce the concentration of IgE in serum or reduce the binding of IgE to the surface receptor of mast cells. In addition, the strain may inhibit the level of MCPT-1 in serum.

[0034] Therefore, the strain according to a specific embodiment can be effectively used for treating allergic diseases.

[0035] On the other hand, there is provided a lysate, culture broth, extract of the culture broth, or a mixture thereof derived from a Lactiplantibacilus plantarum strain.

[0036] In this specification, the term "culture broth" may be used interchangeably with "culture supernatant", "culture supernatant liquor", "conditioned culture broth", or "conditioned medium", and may refer to the entire medium including the strain, its metabolites, and additional nutrients obtained by culturing the strain in a medium capable of supplying nutrients for the Lactiplantibacillus strain to grow and survive in vitro for a certain period of time. The culture is a product obtained by culturing a probiotic strain in a known medium, and the product may or may not include the strain itself. The medium may be selected from known liquid media or solid media, for example, it may be MRS liquid medium, GAM liquid medium, MRS agar medium, GAM agar medium, and BL agar medium, but is not limited thereto.

[0037] The culture solution may include the culture solution itself obtained by culturing the strain, its concentrate or freeze-dried product, or the culture supernatant obtained by removing the strain from the culture solution, its concentrate or freeze-dried product.

[0038] In this specification, the term "lysate" may be used interchangeably with "solubilizate", which refers to a solution or suspension of disrupted microbial cells such as Lactiplantibacillus plantarum in an aqueous medium. The cell lysate includes, for example, macromolecules such as DNA, RNA, proteins, peptides, carbohydrates, and lipids, etc. and / or small molecules such as amino acids, sugars, fatty acids, etc. or fractions thereof. Additionally, the solubilizate contains cell debris, the structure of which may be smooth or granular.

[0039] In this specification, the term "culture solution extract" refers to a substance extracted from the culture solution or its concentrate, and may include an extract solution, a diluted or concentrated solution of the extract solution, a dried product obtained by drying the extract solution, or its crude product or purified product, as well as fractions obtained by fractionating the same.

[0040] In a specific embodiment, the culture solution may be obtained by culturing Lactiplantibacillus plantarum in a suitable medium (e.g., MRS plate medium) at any temperature above 10 °C or below 40 °C for a certain period of time, such as 4 hours to 50 hours.

[0041] In a specific embodiment, those of ordinary skill in the art can appropriately select or modify the medium and culture conditions for culturing the Lactiplantibacillus plantarum.

[0042] On the other hand, there is provided a composition comprising a lysate, a culture solution, an extract of the culture solution, or a mixture thereof derived from a Lactiplantibacilus plantarum strain.

[0043] In a specific embodiment, the composition may further comprise an additive selected from the group consisting of preservatives, dyes, emulsifiers, sweeteners, stabilizers, flavor enhancers, flavoring agents, and acidulants.

[0044] In a specific embodiment, the composition may be one or more dosage forms selected from the group consisting of solutions, emulsions, viscous mixtures, powders, granules, tablets, and capsules.

[0045] In a specific embodiment, the composition may have anti-allergic activity.

[0046] In a specific embodiment, the anti-allergy activity may have the activity of inhibiting Th2-mediated cytokine production, inhibiting the degranulation activity of mast cells, inhibiting the Ig level in serum or the MCPT-1 level in serum, and inhibiting food allergy. Specifically, the inhibition of mast cell degranulation activity may be the inhibition of the secretion of β-hexosaminidase from activated mast cells.

[0047] In a specific embodiment, the Th2-mediated cytokine may be one or more selected from the group consisting of IL-4, IL-5, IL-6, IL-10, and IL-13.

[0048] In a specific embodiment, the composition may reduce the concentration of IgE in serum or reduce the binding of IgE to the surface receptor of mast cells. In addition, the strain may inhibit the level of MCPT-1 in serum.

[0049] According to a specific embodiment, the anti-allergy activity may include the activity of inhibiting mast cell degranulation by 50% to 70% when treating the supernatant of Lactobacillus plantarum GBCC_F0070 strain in mast cell RBL-2H3.

[0050] According to a specific embodiment, the anti-allergy activity may include the activity of inhibiting ear swelling by 30% to 50% when treating the supernatant of Lactobacillus plantarum GBCC_F0070 strain in an atopic dermatitis animal model induced by MC903.

[0051] According to a specific embodiment, the anti-allergy activity may include the activity of reducing the IgE concentration in serum by 50% to 70% or reducing the MCPT-1 concentration in serum by 50% to 70% when treating the supernatant of Lactobacillus plantarum GBCC_F0070 strain in an atopic dermatitis animal model induced by MC903.

[0052] On the other hand, provided is the use of a Lactobacillus plantarum, a lysate, a culture solution, or an extract of the strain in improving, preventing, or treating a disease.

[0053] In a specific embodiment, the use may include preventing, improving, or treating a disease or disorder mediated by IgE (ImmunoglobulinE) or not mediated by IgE. Specifically, the disease mediated by IgE (Immunoglobulin E) or not mediated by IgE may be an allergic disease.

[0054] In this specification, "allergy disease" refers to a disease caused by an allergic reaction, which is an excessive immune reaction in the body to external antigens. Specifically, it can be one or more diseases selected from the group consisting of edema, anaphylaxis, allergic rhinitis, asthma, allergic conjunctivitis, allergic dermatitis, atopic dermatitis, contact dermatitis, urticaria, chronic spontaneous urticaria, pruritus, anaphylactic shock, insect allergy, food allergy, drug allergy, and respiratory allergy, but is not limited thereto.

[0055] The development process of the allergy disease is generally divided into an early reaction and a late reaction. In the early reaction, when an antibody (IgE) is generated by antigen stimulation and then binds to the high-affinity receptor (FcεRI, high affinity receptor) on the surface of mast cells, when the antigen re-invades, it binds to the antibody (IgE) bound to FcεRI, causing a signal to be rapidly transmitted into the mast cells, resulting in the secretion of inflammatory chemical mediators (cytokines, histamine, leukotrienes) that cause allergies.

[0056] In the late reaction, Th2-type cytokines (IL-4, IL-5, IL-6, IL-10, IL-13) induce the production of chemokines (monocyte chemoattractant protein-1, eotaxin-1, RANTES) in tissue fibroblasts or epithelial cells, thereby causing inflammatory cells, especially eosinophils, to migrate to the site of the allergic reaction. As a result, most allergic symptoms are caused by inflammatory reaction mediators such as histamine secreted by mast cells or eosinophils, leading to clinical symptoms such as itching, edema, congestion, cough, and excessive phlegm caused by vasodilation of capillaries around epithelial cells such as the respiratory tract and skin, influx of immune cells, muscle contraction, and increased mucus secretion.

[0057] In particular, among Th2-type cytokines, IL-4 is produced by T cells and mast cells and plays roles such as regulating B cell class switching and promoting IgE production. IL-5 affects the production, activation, and survival of eosinophils, and eosinophils are activated by IgE. Therefore, the activities of IL-4, IL-5, and eosinophils are associated with increased IgE.

[0058] In a specific embodiment, the allergic disease may include allergic reactions caused by indoor dust, fungi, mites, or animal hair, skin, or excrement.

[0059] In a specific embodiment, the allergic disease may be a disease or disorder mediated or not mediated by IgE (Immunoglobulin E), or a condition associated with allergic diseases.

[0060] In this specification, the term "treat" may refer to curing inflammation, bacterial infections, etc. in a shorter time compared to natural healing. The treatment may include improving and / or alleviating inflammation or bacterial infections. Additionally, the treatment may mean curing and / or restoring the symptoms induced by inflammation or bacterial infections.

[0061] In this specification, the term "comprising... as an active ingredient" means adding the strain of this specification, the lysate of the strain, the culture solution, or an extract of the culture solution to such an extent that the above effects can be achieved, and it means including adding various components as auxiliary components for drug delivery, stabilization, etc. to formulate into various forms.

[0062] In a specific embodiment, based on the total weight of the composition, the composition may include 0.001 wt% to 80 wt% of Lactobacillus plantarum strains. Additionally, the dosage of the Lactobacillus plantarum strains may be 0.01 mg to 10000 mg, 0.1 mg to 1000 mg, 1 mg to 100 mg, 0.01 mg to 1000 mg, 0.01 mg to 100 mg, 0.01 mg to 10 mg, or 0.01 mg to 1 mg. The strain is included in the composition at a therapeutically effective amount or a nutritionally effective concentration. For example, the content of the strain may be 10 3 to 10 16 CFU / g, 10 3 to 10 15 CFU / g, 10 3 to 10 14 CFU / g, 10 3 to 10 13 CFU / g, 103 to 10 12 CFU / g, 10 4 to 10 16 CFU / g, 10 4 to 10 15 CFU / g, 10 4 to 10 14 CFU / g, 10 4 to 10 13 CFU / g, 10 4 to 10 12 CFU / g, 10 5 to 10 16 CFU / g, 10 5 to 10 15 CFU / g, 10 5 to 10 14 CFU / g, 10 5 to 10 13 CFU / g, 10 5 to 10 12 CFU / g, 10 6 to 10 13 CFU / g, 10 6 to 10 12 CFU / g, 10 7 to 10 13 CFU / g, 10 7 to 10 12 CFU / g, 10 8 to 10 13 CFU / g or 10 8 to 10 12 CFU / g, or may be included in the composition as a culture of an equivalent amount of live or dead bacteria. Specifically, for adult patients, 1×10 3 CFU / g to 1×10 16Live or dead bacteria at CFU / g. However, depending on factors such as the formulation method, administration method, patient's age, weight, gender, pathological condition, diet, administration time, administration route, excretion rate, and sensitivity to the drug, the administration amount can be prescribed in various ways, and those of ordinary skill in the art can appropriately adjust the administration amount by considering these factors. The number of administrations can be once a month, once every two weeks, once a week, once a day, twice a day, or three times a day. Specifically, it can be administered once or two or more times within the clinically acceptable range of side effects, and the administration site can be one, two, or more. For animals other than humans, the administration amount per kg (body weight) is the same as that for humans, or, for example, it is administered in an amount obtained by converting the above administration amount through the volume ratio (such as the average value) of organs (such as the heart) between the target animal and humans. Possible administration routes can include oral, sublingual, parenteral (such as subcutaneous, intradermal, intravenous, intra-arterial, intramuscular, intraperitoneal, intrathecal, intra-articular, intrasynovial, intrasternal, or intralesional), rectal, topical (including transdermal), inhalation, and injection, or the insertion of an implantable device or substance. As the animals to be treated according to a specific embodiment, it can include humans and other target mammals, specifically including humans, monkeys, mice, rats, rabbits, sheep, cows, dogs, horses, pigs, etc. According to one embodiment, the composition includes dead and dried strains, and can be administered once at 1 g to 10 g, 0.5 g to 1.5 g, 2.5 g to 3.5 g, or 4.5 g to 5.5 g, and can be administered one to three times a day.

[0063] On the other hand, a health food composition for preventing or improving allergic diseases is provided, which includes Lactobacillus plantarum strains, lysates, culture broths, or mixtures thereof derived from the strains as active ingredients.

[0064] The "strains", "anti-allergic activity", and "allergic diseases" are as defined above.

[0065] In a specific embodiment, the health food composition can have anti-allergic activity.

[0066] In a specific embodiment, the anti-allergic activity can have the activity of inhibiting Th2-mediated cytokine production, inhibiting the degranulation activity of mast cells, inhibiting the Ig level in serum, or inhibiting the MCPT-1 level in serum. Specifically, the inhibition of mast cell degranulation activity means that it can inhibit the secretion of β-hexosaminidase from activated mast cells.

[0067] In a specific embodiment, the Th2-mediated cytokines can be one or more selected from the group consisting of IL-4, IL-5, IL-6, IL-10, and IL-13.

[0068] In a specific embodiment, the health food composition can reduce the concentration of IgE in the serum or reduce the binding of IgE to the surface receptor of mast cells. In addition, the strain can inhibit the level of MCPT-1 in the serum.

[0069] Preferably, the health food composition is made into any dosage form selected from powders, granules, pills, tablets, capsules, candies, syrups, and beverages, but is not limited thereto. The health food composition of the present invention can be manufactured by directly adding the active ingredient or mixing it with other foods or food ingredients, and can be appropriately manufactured according to conventional methods. Examples of foods to which the extract can be added can be any form selected from caramels, meats, sausages, breads, chocolates, confectioneries, snacks, cookies, pizzas, ramen, other noodles, chewing gums, dairy products (including ice creams, infant formula milks, milks, yogurts, cheeses, fermented milks, milk powders, etc.), various soups, beverages, teas, drinks, alcoholic beverages, and multivitamins, and include all health foods in the conventional sense. That is, there is no particular limitation on the type of the food. The health food composition can contain various nutrients, vitamins, minerals (electrolytes), synthetic and natural flavoring agents, coloring agents, and thickening agents (such as cheeses and chocolates), pectic acids and their salts, alginic acids and their salts, organic acids, protective colloid thickening agents, pH regulators, stabilizers, preservatives, glycerol, alcohol, carbonating agents used in carbonated beverages, etc. In addition, it can also contain the pulp used for manufacturing natural fruit and vegetable beverages. The above components can be used alone or in combination.

[0070] In a specific embodiment, the improvement of the allergic disease can include alleviating the immune hypersensitivity reaction or improving the skin condition caused by the immune hypersensitivity reaction.

[0071] In a specific embodiment, the improvement of the allergic disease can include inhibiting the degranulation activity of mast cells or inhibiting the levels of IgE or MCPT-1 in the serum.

[0072] In a specific embodiment, the allergic disease can be selected from the group consisting of edema, anaphylaxis, allergic rhinitis, asthma, allergic conjunctivitis, allergic dermatitis, atopic dermatitis, contact dermatitis, urticaria, chronic spontaneous urticaria, pruritus, anaphylactic shock, insect allergy, food allergy, drug allergy, and respiratory allergy.

[0073] In a specific embodiment, the allergic disease may include an allergic reaction caused by indoor dust, fungi, mites, or animal hair, skin, or excrement.

[0074] The health food composition may use the strain or its culture solution alone, or in combination with other foods or food ingredients, and may be appropriately used according to conventional methods. The mixing amount of the active ingredient can be appropriately determined according to the purpose of use (prevention, health care, or treatment). Generally, when manufacturing food or beverage, the composition of the present specification can be added in an amount of 15 parts by weight or less based on the raw materials. There is no particular limitation on the type of the health food. Among the types of health foods, the beverage composition may contain various flavoring agents or natural carbohydrates, etc. as additional ingredients like ordinary beverages. The natural carbohydrates are monosaccharides such as glucose and fructose; disaccharides such as maltose and sucrose; polysaccharides such as dextrin and cyclodextrin; sugar alcohols such as xylitol, sorbitol, and erythritol. As sweeteners, natural sweeteners such as thaumatin and stevia extract, or synthetic sweeteners such as saccharin and aspartame can be used. The health food composition may also contain nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH regulators, stabilizers, preservatives, glycerol, alcohol, and carbonating agents used in carbonated beverages or combinations thereof. The health food composition may also contain the pulp for manufacturing natural fruit juices, fruit juice beverages, vegetable beverages, or combinations thereof.

[0075] On the other hand, a pharmaceutical composition for treating or preventing allergic diseases is provided, which comprises a Lactobacillus plantarum strain, a lysate, a culture solution, or a mixture derived from the strain as an active ingredient.

[0076] The "strain", "anti-allergic activity", and "allergic disease" are as defined above.

[0077] In a specific embodiment, the pharmaceutical composition may have anti-allergic activity.

[0078] In a specific embodiment, the anti-allergic activity may have the activity of inhibiting Th2-mediated cytokine production, inhibiting the degranulation activity of mast cells, inhibiting the Ig level in serum, or inhibiting the MCPT-1 level in serum. Specifically, the inhibition of mast cell degranulation activity means that it can inhibit the secretion of β-hexosaminidase from activated mast cells.

[0079] In a specific embodiment, the Th2-mediated cytokine may be one or more selected from the group consisting of IL-4, IL-5, IL-6, IL-10, and IL-13.

[0080] In a specific embodiment, the pharmaceutical composition can reduce the concentration of IgE in serum or reduce the binding of IgE to the surface receptor of mast cells. In addition, the strain can inhibit the level of MCPT-1 in serum.

[0081] In a specific embodiment, the allergic disease can be selected from the group consisting of edema, anaphylaxis, allergic rhinitis, asthma, allergic conjunctivitis, allergic dermatitis, atopic dermatitis, contact dermatitis, urticaria, chronic spontaneous urticaria, pruritus, anaphylactic shock, insect allergy, food allergy, drug allergy, and respiratory allergy.

[0082] In a specific embodiment, the allergic disease can include allergic reactions caused by indoor dust, fungi, mites, or animal hair, skin, or excrement.

[0083] The pharmaceutical composition may further include a pharmaceutically acceptable diluent or carrier. The diluent can be lactose, corn starch, soybean oil, microcrystalline cellulose, or mannitol, and the lubricant can be magnesium stearate, talc, or a combination thereof. The carrier can be an excipient, a disintegrant, a binder, a lubricant, or a combination thereof. The excipient can be microcrystalline cellulose, lactose, low-substituted hydroxypropyl cellulose, or a combination thereof. The disintegrant can be calcium carboxymethyl cellulose, sodium starch glycolate, anhydrous monobasic calcium phosphate, or a combination thereof. The binder can be polyvinylpyrrolidone, low-substituted hydroxypropyl cellulose, hydroxypropyl cellulose, or a combination thereof. The lubricant can be magnesium stearate, silica, talc, or a combination thereof.

[0084] The pharmaceutical composition can be formulated into an oral or parenteral dosage form. Specifically, it can be formulated for oral, rectal, intravenous, intra-vascular, intratumoral, subcutaneous, intradermal, or intraperitoneal delivery. More specifically, the oral administration dosage form can be granules, powders, liquids, tablets, capsules, dry syrups, or a combination thereof, and the parenteral administration dosage form can be an injection.

[0085] On the other hand, a cosmetic composition for preventing or improving allergic diseases is provided, which includes a Lactobacillus plantarum strain, a lysate, a culture solution, or a mixture thereof derived from the strain as an active ingredient.

[0086] The "strain", "anti-allergic activity", and "allergic disease" are as defined above.

[0087] In a specific embodiment, the cosmetic composition may have anti-allergic activity.

[0088] The cosmetic composition may have a cosmetic dosage form such as a soft lotion, a nutritive lotion, a massage cream, a nutritive cream, a serum, a mask, a gel, an ampoule, or a skin-adhesive type.

[0089] In addition to the above composition, the ingredients contained in the cosmetic composition may further include ingredients commonly used in cosmetic compositions as active ingredients, for example, may include conventional auxiliaries and carriers, such as stabilizers, solubilizers, vitamins, pigments, and fragrances.

[0090] On the other hand, there is provided a method for preventing, improving, or treating the condition of an individual, which comprises the step of treating or administering an effective amount of the composition to an individual in need thereof.

[0091] In a specific embodiment, the condition of the individual may be a disease or disorder mediated by IgE (Immunoglobulin E) or not mediated by IgE, or a condition related to allergic diseases.

[0092] In the present specification, the term "administer" refers to physically introducing the composition into the body of an object using any of a variety of methods and delivery systems known to those of ordinary skill in the art. The administration can be carried out by methods known in the art. The administration can be directly administered to an individual in any manner through a route, including, for example, intravenous, intramuscular, oral, transdermal, mucosal, intranasal, intratracheal, or subcutaneous administration. The administration can be systemic or local administration.

[0093] The individual may be a mammal, such as a human, a cow, a horse, a pig, a dog, a sheep, a goat, or a cat. The individual may be an individual in need of improving the effect of a condition related to inflammation or a condition related to bacterial infection.

[0094] The administration may be administering the composition according to a specific embodiment to each individual in an amount of 0.00001 mg to 1000 mg per day. For example, it may be administered in an amount of 0.00001 mg to 500 mg, 0.00001 mg to 100 mg, 0.00001 mg to 50 mg, 0.00001 mg to 25 mg, 1 mg to 1000 mg, 1 mg to 500 mg, 1 mg to 100 mg, 1 mg to 50 mg, 1 mg to 25 mg, 5 mg to 1000 mg, 5 mg to 500 mg, 5 mg to 100 mg, 5 mg to 50 mg, 5 mg to 25 mg, 10 mg to 1000 mg, 10 mg to 500 mg, 10 mg to 100 mg, 10 mg to 50 mg, or 10 mg to 25 mg. However, depending on factors such as the formulation method, administration method, the patient's age, weight, gender, pathological condition, diet, administration time, administration route, excretion rate, and sensitivity to the drug, the dosage can be prescribed in various ways, and those of ordinary skill in the art can appropriately adjust the dosage by considering these factors. The number of administrations can be once a day or two or more times within a clinically acceptable range of side effects. The administration site can be one, two, or more, and it can be administered daily or at intervals of 2 to 5 days, and the total number of days of administration for one treatment is 1 to 30 days. If necessary, the same treatment can be repeated after an appropriate time. For animals other than humans, the dosage per kg is the same as that for humans, or, for example, it is administered in an amount obtained by converting the above dosage through the volume ratio (such as the average value) of organs (such as the heart) between the target animal and humans. Beneficial effects

[0095] The new strain according to one aspect and the lysate, culture solution, culture solution extract, or a mixture thereof derived therefrom have the effect of being effectively used for preventing, improving, or treating allergic diseases. Brief description of the drawings

[0096] Figure 1 is the genomic map of the GBCC_F0070 strain.

[0097] Figure 2 is a graph showing the effect of the GBCC_F0070 culture supernatant on the degranulation inhibition of RBL-2H3 mast cells.

[0098] Figure 3 is a graph showing the effect of GBCC_F0070 on ear swelling inhibition in a mouse model of atopic dermatitis induced by MC903.

[0099] Figure 4 is a graph showing the effect of GBCC_F0070 on reducing IgE in serum in a mouse model of atopic dermatitis induced by MC903.

[0100] Figure 5 It is a graph showing the effect of reducing MCPT-1 in the serum of a mouse model of atopic dermatitis induced by MC903 by GBCC_F0070.

[0101] Figure 6 It is a graph showing the inhibitory effect on IgE-mediated allergic reactions in a passive cutaneous anaphylaxis animal model in the group administered with Lactobacillus plantarum GBCC_F0070 strain.

[0102] Figure 7 It is a graph showing the effect of improving allergic symptoms in a food allergy animal model in the group administered with Lactobacillus plantarum GBCC_F0070 strain.

[0103] Figure 8 It is a graph showing the effect of reducing total IgE in the serum in a food allergy animal model in the group administered with Lactobacillus plantarum GBCC_F0070 strain.

[0104] Figure 9 It is a graph showing the effect of reducing inflammatory cells in bronchoalveolar lavage fluid (BALF) in an ovalbumin-induced asthma animal model in the group administered with Lactobacillus plantarum GBCC_F0070 strain.

[0105] Figure 10 It is a graph showing the effect of reducing total IgE in the serum in an ovalbumin-induced asthma animal model in the group administered with Lactobacillus plantarum GBCC_F0070 strain.

[0106] Figure 11 It is a graph showing the inhibitory effect on Th2 cytokines in the serum in an ovalbumin-induced asthma animal model in the group administered with Lactobacillus plantarum GBCC_F0070 strain.

[0107] Figure 12 It is a graph showing the inhibitory effect on Th2 cytokines (IL-4, IL-5, IL-13) in the nasal mucosa in an ovalbumin-induced rhinitis animal model in the group administered with Lactobacillus plantarum GBCC_F0070 strain.

[0108] Figure 13 It is a graph showing the effect of reducing total IgE and ovalbumin-specific IgE in the serum in an ovalbumin-induced rhinitis animal model in the group administered with Lactobacillus plantarum GBCC_F0070 strain. Detailed implementation manners

[0109] In the following, preferred embodiments are presented to assist in understanding the present invention. However, the following embodiments are provided only to make the present invention more easily understood, and the content of the present invention is not limited to the following embodiments. These embodiments can be variously changed, and these embodiments are not limited to the embodiments disclosed below and can be implemented in various forms.

[0111] Example 1. Isolation and identification of Lactiplantibacillus plantarum strains

[0112] 1.1 Isolation of strains

[0113] The Lactiplantibacillus plantarum strains of the present invention were isolated from pickled radish pickles at home. After pickling, the pickled radish pickles after 23 days were spread on MRS (deMan, Rogosa, Sharpe) plate medium as a lactic acid bacteria medium by the method of serial dilution at 1 / 10 in PBS buffer. After culturing for 2 days in an anaerobic chamber at 37 °C, the colonies were observed, and the single colonies were subcultured onto a new MRS plate medium for pure isolation. The identification of the pure isolated strains was confirmed by PCR and Sanger sequencing of the 16S rRNA gene. To preserve the strains for a long time, the strains were cultured in liquid, and 30% glycerol was added at a ratio of 1:1 to the cells at the late exponential growth phase, and then stored in an ultra-low temperature freezer at -70 °C. Pure isolation and identification were also carried out in MRS plate medium and liquid medium. When cultured in MRS solid medium for 48 hours, large, round white colonies with a smooth surface were formed.

[0114] Through the above method, the Lactiplantibacillus plantarum GBCC_F0070 strain (hereinafter also referred to as "GBCC_F0070") was finally selected.

[0116] 1.2 Molecular biological identification of the selected strains

[0117] The Lactiplantibacillus plantarum GBCC_F0070 strain was identified based on the sequence similarity of the 16S rRNA gene and the recA gene. DNA was extracted from the bacterial culture using SPIN Soil Kit (MP Bio, 6560200), and PCR was performed using the primer sequences shown in Table 1.

[0118]

Table 1

[0119] The sequence generated using the 27F and 1492R primers for PCR amplification of the 16S rRNA gene of bacteria is shown in SEQ ID NO: 1. The 16S rRNA sequence of GBCC_F0070 showed a sequence similarity of 98.5% or higher with 13 species of the Lactiplantibacillus sp. including the type strain of Lactiplantibacillus plantarum, and thus it was confirmed that GBCC_F0070 corresponded to one of these 13 species strains (Table 2).

[0120]

Table 2

[0121] Before the change of the strain name, Lactiplantibacillus plantarum was named Lactobacillus plantarum. In addition, in addition to the 16S rRNA gene, more accurate identification was carried out based on the sequence similarity of the recA gene widely used as a taxonomic biomarker for bacteria. As a result, it was confirmed that GBCC_F0070 was the strain closest to the species of Lactiplantibacillus plantarum (Table 3).

[0122]

Table 3

[0123] The recA gene sequence of GBCC_F0070 is shown in SEQ ID NO: 2.

[0124] Example 2. Genome and comparative genome analysis of Lactiplantibacillus plantarum strains

[0125] To investigate the species identification and characteristics based on the genome of the GBCC_F0070 strain, the next-generation sequencing technology (NGS) and bioinformatics technology were used to completely interpret the genome sequence of this strain, and the functions of the genes contained in the genome were inferred.

[0126] The genomic interpretation results confirmed that GBCC_F0070 has a circular chromosome of 3,259,176 bp and 6 plasmids, and also confirmed the presence of 3,140 CDSs, 16 rRNA genes and 66 tRNA genes in GBCC_F0070( Figure 1 , Table 4).

[0127]

Table 4

[0128] The average nucleotide identity (ANI) of the genomes of the strains of the same species published in GenBank and the genome of the GBCC_F0070 strain was calculated using the Jspecies program. The results showed that the sequence similarity with the closest strain was 99.16%. Thus, GBCC_F0070 is a novel strain that has never been reported before (Table 5).

[0129]

Table 5

[0130] Therefore, the inventors named the GBCC_F0070 strain "Lactiplantibacillus plantarum GBCC_F0070" (Accession No.: KCTC 15051BP), and deposited it at the Korean Collection for Type Cultures (KCTC) of the Korea Research Institute of Bioscience and Biotechnology on August 16, 2022.

[0132] Experimental Example 1. Inhibitory effect of the culture supernatant of Lactiplantibacillus plantarum GBCC_F0070 on degranulation of RBL-2H3 mast cells

[0133] To demonstrate the therapeutic activity against IgE-mediated allergy, the degranulation inhibitory efficacy of mast cells was confirmed. The culture supernatants of various Lactiplantibacillus strains including the GBCC_F0070 strain of Lactiplantibacillus plantarum were separately treated with activated mast cells that induced allergic reactions, and the inhibitory effect on degranulation was screened by measuring the secretion of β-hexosaminidase. β-Hexosaminidase is stored in the secretory granules of mast cells and histamine is secreted extracellularly due to allergic reactions, so it can be used as an indicator factor for mast cell degranulation.

[0134] Suspend RBL-2H3 cells in 15% FBS and MEM medium in a 24-well plate, then seed 1.5×10 5 cells per well and culture with 20 ng / mL of DNP-IgE for about 12 hours to sensitize. Wash the cells once with PIPES buffer (25 mM PIPES, 119 mM NaCl, 5 mM KCl, 0.4 mM MgCl 2 , 1 mM CaCl 2 , 5.6 mM glucose, 0.1% BSA), then dilute the culture supernatants of various Lactobacillus plantarum strains to 1:15 in PIPES buffer and pretreat for 2 hours in each well. The culture supernatant is the supernatant obtained by culturing Lactobacillus plantarum strains in MRS medium, centrifuging to precipitate the strains and only obtaining the supernatant, and then filtering through a 0.22 μm filter. Then add 25 ng / mL of DNP-HSA (antigen) to stimulate for 15 minutes, and then cool on ice for 5 minutes to terminate the reaction. Transfer the supernatant to an e-tube, wash once with PIPES buffer, and then lyse the cells remaining on the plate with 0.1% triton X-100. Mix 30 μL of the supernatant and the lysed cell sample with substrate buffer (1 mM 4-nitrophenyl N-acetyl-β-D-glucosamine, 0.1 M sodium citrate) in a 96-well plate respectively, and react at 37 °C for 1 hour. After adding 0.1 M carbonate buffer to terminate the reaction, measure the absorbance at a wavelength of 405 nm using a microplate reader, as Figure 2 shown.

[0135] Figure 2 is a graph showing the inhibitory effect of the culture supernatant of GBCC_F0070 on the degranulation of RBL-2H3 mast cells.

[0136] As Figure 2 shown, according to the results of screening the culture supernatants of 26 different Lactobacillus plantarum strains, it has been confirmed that the culture supernatant of Lactobacillus plantarum strain GBCC_F0070 shows inhibition of about 61% of mast cell degranulation and is the most effective in inhibiting IgE-mediated allergic reactions.

[0138] Experimental Example 2. Ear swelling inhibitory effect of the group administered with Lactobacillus plantarum strain GBCC_F0070 in an atopic dermatitis animal model induced by MC903

[0139] In an atopic dermatitis animal model induced by MC903, confirm the ear swelling inhibitory effect of Lactobacillus plantarum strain GBCC_F0070.

[0140] Six-week-old BALB / c mice were obtained. After a one-week acclimation period, the freeze-dried Lactobacillus plantarum GBCC_F0070 strain was suspended in D-PBS and orally administered at a dose of 2×10 9 CFU per mouse per day from 7 days before the induction of atopic dermatitis until the end of the experiment. To induce atopic dermatitis, after calibrating the mice, 20 μL of 2 nmol of MC903 (calcipotriol, C 27 H 4 0O 3 ) dissolved in ethanol was applied to both ears of the disease induction group daily for 12 consecutive days, while only ethanol was applied to the normal group. To confirm the degree of ear swelling caused by the induction of atopic dermatitis, the thickness of the mice's ears was measured at the same time every day, as Figure 3 shown. At this time, the change in ear swelling was calculated by using the value obtained by subtracting the ear thickness measured before disease induction from the ear thickness measured daily after disease induction.

[0141] Figure 3 is a graph showing the inhibitory effect of GBCC_F0070 on ear swelling in a mouse model of MC903-induced atopic dermatitis.

[0142] As Figure 3 shown, in the MC903-induced atopic dermatitis model, compared with the control group (MC903), an approximately 34% inhibitory effect on ear swelling was confirmed in the experimental group (MC903 + GBCC_F0070) administered the Lactobacillus plantarum GBCC_F0070 strain.

[0144] Experimental Example 3. Effect of reducing total IgE in serum in an animal model of MC903-induced atopic dermatitis in the group administered the Lactobacillus plantarum GBCC_F0070 strain

[0145] The overexpression of IgE is the most representative marker of atopic dermatitis. To confirm the total IgE concentration in serum from the MC903-induced atopic dermatitis animal model in the above Experimental Example 3, measurement was performed using a mouse IgE ELISA kit (Biolegend #432404).

[0146] The capturing antibody diluted with coating buffer was reacted overnight at 4 °C on microwell plates, then washed four times with washing buffer and blocked with assay diluent for 1 hour at room temperature. After washing four times, standard samples and serum samples diluted 1:1000 were added and reacted at room temperature for 2 hours, then washed four times, and detection antibody was added to react for 1 hour at room temperature. After washing four times, avidin-HRP was added to react for 30 minutes at room temperature, then washed five times, TMB substrate solution was added and incubated in the dark for 20 minutes, and reacted at room temperature. Stop solution was added to each well to terminate the color reaction, and the absorbance was measured at 450 nm using an ELISA reader, as Figure 4 shown.

[0147] Figure 4 is a graph showing the effect of GBCC_F0070 on reducing IgE in the serum of a mouse model of MC903-induced atopic dermatitis.

[0148] As Figure 4 shown, compared with the normal group, the total IgE concentration in the serum of the control group (MC903) increased sharply, but in the experimental group (MC903 + GBCC_F0070) administered with Lactobacillus plantarum GBCC_F0070 strain, a reduction effect of about 62% in the total IgE concentration in the serum was confirmed.

[0150] Experimental Example 4. Effect of Lactobacillus plantarum GBCC_F0070 strain administration group on reducing MCPT-1 in the serum of a MC903-induced atopic dermatitis animal model

[0151] To confirm the concentration of mast cell protease-1 (MCPT-1) in the serum from the MC903-induced atopic dermatitis animal model in Experimental Example 2, measurement was performed using a mouse MCPT-1 (mMCP-1) ELISA kit (Invitrogen #88-7503).

[0152] The capture antibody diluted with coating buffer was reacted overnight on a microplate at 4°C, then washed three times with washing buffer and blocked with ELISA / ELISPOT diluent for 1 hour at room temperature. After washing three times, standard samples and serum samples diluted 1:100 were added, and the reaction was carried out at room temperature for 2 hours. Then, it was washed three times, and a detection antibody was added to react for 1 hour at room temperature. After washing three times, avidin-HRP was added to react for 30 minutes at room temperature, then washed five times, TMB substrate solution was added to react for 15 minutes in the dark, and the reaction was carried out at room temperature. A reaction termination solution was added to each well to terminate the color reaction, and the absorbance was measured at 450 nm using an ELISA reader, as Figure 5 shown.

[0153] Figure 5 is a graph showing the effect of reducing MCPT-1 in the serum of a mouse model of atopic dermatitis induced by MC903 by Lactobacillus plantarum GBCC_F0070.

[0154] As Figure 5 shown, compared with the normal group, the concentration of MCPT-1 in the serum of the control group (MC903) increased sharply, but in the experimental group (MC903+GBCC_F0070) administered with Lactobacillus plantarum GBCC_F0070 strain, a reduction effect of about 57% in the concentration of MCPT-1 in the serum was confirmed.

[0156] Experimental Example 5. Inhibitory effect of Lactobacillus plantarum GBCC_F0070 strain-administered group on IgE-mediated allergic reaction in a passive cutaneous anaphylaxis animal model

[0157] The inhibitory effect of Lactobacillus plantarum GBCC_F0070 strain on allergic reaction was confirmed in an animal model of passive cutaneous anaphylaxis (PCA) caused by anti-DNP IgE.

[0158] Four-week-old BALB / c mice were obtained. After a one-week acclimation period, the freeze-dried Lactobacillus plantarum GBCC_F0070 strain was suspended in D-PBS, and from 2 days before IgE sensitization until the end of the experiment, 1×10 per mouse was administered daily 10The dose of CFU was administered orally. As a positive control group, dexamethasone (Dexa, 10 mg / kg) was used. To induce a passive cutaneous anaphylaxis allergic reaction, 20 ng of anti-DNP IgE was locally injected into one ear of the mice. Approximately 24 hours later, DNP-HSA was diluted to a concentration of 0.5 mg / mL in Evans blue staining solution at 5 mg / mL, and then 200 μL was administered via the tail vein. 30 minutes later, the mice were sacrificed, and the ears were removed and placed in 700 μL of formamide. After that, the staining drug infiltrated into the ear tissue was extracted at 63 °C for 12 hours, and then the absorbance of the supernatant was measured at a wavelength of 620 nm. The quantification of the exuded Evans blue was measured using a standard calibration curve, and the results are as Figure 6 shown.

[0159] Figure 6 is a graph showing the inhibitory effect of the Lactobacillus plantarum GBCC_F0070 strain administration group on IgE-mediated allergic reactions in an animal model of passive cutaneous anaphylaxis.

[0160] As Figure 6 shown, in a passive cutaneous anaphylaxis allergic reaction model induced by anti-DNP IgE, compared with the negative control group (Ag), in the experimental group (Ag+GBCC_F0070) administered with the Lactobacillus plantarum GBCC_F0070 strain, an anti-allergic effect of inhibiting approximately 24% of the allergic reaction was confirmed.

[0162] Experimental Example 6. Effect of the Lactobacillus plantarum GBCC_F0070 strain administration group on improving allergic symptoms in an animal model of food allergy

[0163] In an animal model of food allergy induced by ovalbumin (OVA), the effect of the Lactobacillus plantarum GBCC_F0070 strain on improving allergic symptoms was confirmed.

[0164] Five-week-old BALB / c mice were obtained. After a one-week acclimation period, to induce a food allergy reaction, 50 μg of OVA and 1 mg of aluminum hydroxide were mixed and intraperitoneally injected 2 times every two weeks for sensitization. Two weeks after the last sensitization date, 50 mg of ovalbumin was orally administered 5 times every two days to induce an allergic reaction. In addition, to confirm the allergic treatment effect, the freeze-dried Lactobacillus plantarum GBCC_F0070 strain was suspended in D-PBS, and from the start of ovalbumin sensitization until the end of the experiment, it was orally administered to each mouse at a dose of 2×10 9 CFU per day. The stool morphology caused by food allergy was observed macroscopically, and the diarrhea symptoms were scored. The results are shown in Figure 7and Table 6.

[0165] [[Table 6]]

[0166] Figure 7 is a chart showing the improvement effect of the allergic symptoms in the food allergy animal model of the Lactobacillus plantarum GBCC_F0070 strain application group. As Figure 7 shown, compared with the normal group, the diarrhea symptom score of the control group (OVA) induced with food allergy increased significantly, and in the food allergy reaction model caused by ovalbumin, compared with the control group, in the experimental group (OVA+GBCC_F0070) applying the Lactobacillus plantarum GBCC_F0070 strain, an improvement effect of about 26% of the diarrhea symptoms could be confirmed.

[0168] Experimental Example 7. Reduction effect of total IgE in serum in the food allergy animal model of the Lactobacillus plantarum GBCC_F0070 strain application group

[0169] In order to confirm the total IgE concentration in serum from the food allergy animal model induced by ovalbumin, measurement was carried out using a mouse IgE ELISA kit (Biolegend#432404).

[0170] The capture antibody diluted with coating buffer was reacted overnight at 4 °C on a microplate, then washed four times with washing buffer and blocked with assay diluent for 1 hour at room temperature. Washed four times and standard samples and serum samples diluted 1:1000 were added, reacted at room temperature for 2 hours, then washed four times, and detection antibody was added to react for 1 hour at room temperature. Washed four times and avidin-HRP was added to react for 30 minutes at room temperature, then washed five times, TMB substrate solution was added to react for 20 minutes in the dark, and reacted at room temperature. Stop solution was added to each well to terminate the color reaction, and the absorbance was measured at 450 nm with an ELISA reader, as Figure 8 shown.

[0171] Figure 8 is a chart showing the reduction effect of total IgE in serum in the food allergy animal model of the Lactobacillus plantarum GBCC_F0070 strain application group.

[0172] As Figure 8 shown, compared with the normal group, the total IgE concentration in the serum of the control group (OVA) increased sharply, but in the experimental group (OVA+GBCC_F0070) applying the Lactobacillus plantarum GBCC_F0070 strain, a reduction effect of about 21% of the total IgE concentration in serum was confirmed.

[0174] Experimental Example 8. Effect of reducing inflammatory cells in bronchoalveolar lavage fluid (BALF) of the Lactobacillus plantarum GBCC_F0070 strain administration group in an ovalbumin-induced asthma animal model

[0175] In an ovalbumin (OVA)-induced bronchial asthma animal model, the asthma alleviating effect of the Lactobacillus plantarum GBCC_F0070 strain was confirmed.

[0176] Five-week-old BALB / c mice were obtained. After a one-week acclimation period, to prepare an asthma mouse model, 50 μg of OVA and 1 mg of aluminum hydroxide were mixed and intraperitoneally injected twice at two-week intervals for sensitization. One week after the last sensitization day, that is, from the 21st to the 25th, 150 μg of ovalbumin was instilled intranasally under anesthesia so that it was inhaled into the lungs to induce asthma. Twenty-four hours after the last challenge, anesthesia was performed and the bronchi were incised, bronchoalveolar lavage fluid was collected, and the total number of cells was counted. The results are as Figure 9 shown.

[0177] Figure 9 It is a graph showing the effect of reducing inflammatory cells in bronchoalveolar lavage fluid (BALF) of the Lactobacillus plantarum GBCC_F0070 strain administration group in an ovalbumin-induced asthma animal model.

[0178] As Figure 9 shown, compared with the normal group, the inflammatory cells in the bronchoalveolar lavage fluid of the asthma-induced group were significantly increased, and compared with the control group (OVA), in the experimental group (OVA + GBCC_F0070) administered with the Lactobacillus plantarum GBCC_F0070 strain, it was confirmed that the inflammatory cells in the bronchoalveolar lavage fluid were reduced by approximately 64%.

[0180] Experimental Example 9. Effect of reducing total IgE and ovalbumin-specific IgE in serum of the Lactobacillus plantarum GBCC_F0070 strain administration group in an ovalbumin-induced asthma animal model

[0181] To confirm the concentrations of total IgE and ovalbumin (OVA)-specific IgE in serum from an ovalbumin-induced asthma animal model, a mouse IgE ELISA kit (Biolegend #432404) and a mouse OVA-specific IgE ELISA kit (Biolegend #439807) were used for measurement.

[0182] For the measurement of total IgE in serum, the capture antibody diluted with coating buffer was further reacted overnight at 4 °C on a microplate, then washed four times with washing buffer and blocked with assay diluent for 1 hour at room temperature. The plate was washed four times and standard samples and serum samples diluted at 1:500 (total IgE), 1:2 (ovalbumin-specific IgE) were added, reacted at room temperature for 2 hours, then washed four times, and detection antibody was added to react for 1 hour at room temperature. It was washed four times and avidin-HRP was added to react for 30 minutes at room temperature, then washed five times, TMB substrate solution was added to react for 20 minutes in the dark and reacted at room temperature. Termination solution was added to each well to terminate the color reaction, and the absorbance was measured at 450 nm with an ELISA reader, as Figure 10 shown.

[0183] Figure 10 is a graph showing the effect of reducing total IgE in the serum of the group administered with Lactobacillus plantarum strain GBCC_F0070 in an ovalbumin-induced asthma animal model.

[0184] As Figure 10 shown, compared with the normal group, the total IgE concentration in the serum of the control group (OVA) increased sharply, but in the experimental group administered with Lactobacillus plantarum strain GBCC_F0070 (OVA+GBCC_F0070), a reduction effect of about 37% in the total IgE concentration in the serum was confirmed.

[0186] Experimental Example 10. Inhibitory effect of Th2 cytokines in the serum of the group administered with Lactobacillus plantarum strain GBCC_F0070 in an ovalbumin-induced asthma animal model

[0187] To confirm the concentration of IL-5 in the serum from an ovalbumin-induced asthma animal model, measurement was performed using a mouse IL-5 ELISA kit (Biolegend #431204).

[0188] The capture antibody diluted with coating buffer was reacted overnight at 4 °C on a microplate, then washed four times with washing buffer and blocked with assay diluent for 1 hour at room temperature. The plate was washed four times and standard samples and serum samples diluted at 1:5 were added, reacted at room temperature for 2 hours, then washed four times, and detection antibody was added to react for 1 hour at room temperature. It was washed four times and avidin-HRP was added to react for 30 minutes at room temperature, then washed five times, TMB substrate solution was added to react for 20 minutes in the dark and reacted at room temperature. Termination solution was added to each well to terminate the color reaction, and the absorbance was measured at 450 nm with an ELISA reader, as Figure 11 shown.

[0189] Figure 11It is a graph showing the inhibitory effect of Lactobacillus plantarum GBCC_F0070 strain administration group on Th2 cytokines in the serum of an ovalbumin-induced asthma animal model.

[0190] As Figure 11 shown, compared with the normal group, the concentration of IL-5 in the serum of the control group (OVA) increased sharply, but in the experimental group (OVA+GBCC_F0070) administered with Lactobacillus plantarum GBCC_F0070 strain, it was confirmed that the production amount of IL-5 in the serum was significantly reduced by about 88% statistically compared with the control group.

[0191] The above content confirmed that Lactobacillus plantarum GBCC_F0070 strain has an inhibitory effect on Th2 cytokines in the serum, which means that it not only has therapeutic activity against IgE-mediated allergies, but also has therapeutic activity against non-IgE-mediated allergies.

[0193] Experimental Example 11. Inhibitory effect of Lactobacillus plantarum GBCC_F0070 strain administration group on Th2 cytokines (IL-4, IL-5) in the nasal mucosa of an ovalbumin-induced rhinitis animal model

[0194] The rhinitis alleviating effect of Lactobacillus plantarum GBCC_F0070 strain was confirmed in an ovalbumin (OVA, ovalbumin)-induced rhinitis animal model.

[0195] Five-week-old BALB / c mice were obtained. After a one-week acclimation period, to make a rhinitis mouse model, 50 μg of OVA and 2 mg of aluminum hydroxide were mixed and intraperitoneally injected 3 times every other week for sensitization. One week after the last sensitization day, that is, from the 21st to the 26th, 100 μg of ovalbumin was instilled intranasally without anesthesia to induce rhinitis while preventing the antigen from being inhaled into the lungs. 24 hours after the last challenge, the nasal mucosa was extracted and broken to isolate RNA. After synthesizing cDNA, the mRNA expression levels of Th2 cytokines (IL-4, IL-5, IL-13) were confirmed using a QuantStudio 3 real-time PCR instrument, and the results are as Figure 12 shown.

[0196] Figure 12 It is a graph showing the inhibitory effect of Lactobacillus plantarum GBCC_F0070 strain administration group on Th2 cytokines (IL-4, IL-5) in the nasal mucosa of an ovalbumin-induced rhinitis animal model.

[0197] As Figure 12As shown, compared with the normal group, the Th2 cytokines in the nasal mucosa of the rhinitis induction group were significantly increased. Compared with the control group (OVA), in the experimental group (OVA + GBCC_F0070) administered with the Lactobacillus plantarum GBCC_F0070 strain, an approximately 25% inhibitory effect on the IL-4 mRNA expression level was shown to be statistically significant, and a 21% decrease in the IL-5 mRNA expression level was confirmed.

[0198] The above content confirmed that the Lactobacillus plantarum GBCC_F0070 strain has an inhibitory effect on Th2 cytokines in the serum, which means that it has not only therapeutic activity against IgE-mediated allergies but also therapeutic activity against non-IgE-mediated allergies.

[0200] Experimental Example 12. Effect of reducing total IgE and ovalbumin-specific IgE in the serum of the Lactobacillus plantarum GBCC_F0070 strain-administered group in an ovalbumin-induced rhinitis animal model

[0201] To confirm the concentrations of total IgE and ovalbumin (OVA)-specific IgE in the serum from an ovalbumin-induced rhinitis animal model, a mouse IgE ELISA kit (Biolegend #432404) and a mouse OVA-specific IgE ELISA kit (Biolegend #439807) were used for measurement.

[0202] For the measurement of total IgE in the serum, the process of reacting the capture antibody diluted with coating buffer overnight at 4°C on a microplate, then washing four times with washing buffer and blocking for 1 hour at room temperature with assay diluent was further carried out. The plate was washed four times and standard samples and serum samples diluted at 1:500 (total IgE), 1:2 (ovalbumin-specific IgE) were added, reacted at room temperature for 2 hours, then washed four times, and a detection antibody was added to react at room temperature for 1 hour.

[0203] Washed four times and avidin-HRP was added to react at room temperature for 30 minutes, then washed five times, TMB substrate solution was added to react in the dark for 20 minutes, and reacted at room temperature. A termination solution was added to each well to terminate the color reaction, and the absorbance was measured at 450 nm with an ELISA reader, as Figure 13 shown.

[0204] Figure 13 is a chart showing the effect of reducing total IgE and ovalbumin-specific IgE in the serum of the Lactobacillus plantarum GBCC_F0070 strain-administered group in an ovalbumin-induced rhinitis animal model.

[0205] As Figure 13As shown, compared with the normal group, the total IgE concentration in the serum of the control group (OVA) increased sharply. However, in the experimental group administered with the Lactobacillus plantarum GBCC_F0070 strain (OVA + GBCC_F0070), a reduction effect of approximately 14% in the total IgE concentration in the serum was confirmed. In addition, compared with the control group, in the experimental group administered with the Lactobacillus plantarum GBCC_F0070 strain, a reduction effect of approximately 22% in the ovalbumin-specific IgE in the serum was confirmed.

[0207] From the above results, it is indicated that the Lactobacillus plantarum GBCC_F0070 strain can be effectively used for the prevention, improvement, or treatment of atopic dermatitis or allergic diseases.

[0209] The above description of the present invention is for illustrative purposes. Those of ordinary skill in the technical field to which the present invention pertains will understand that the present invention can be easily modified into other specific forms without changing the technical idea or essential features of the present invention. Therefore, the above embodiments and experimental examples should be understood as illustrative in all aspects and not restrictive.

[0210]

Deposit Number

[0211] Name of the depositary institution: Korea Research Institute of Bioscience and Biotechnology

[0212] Deposit Number: KCTC15051BP

[0213] Date of deposit: 20220816

[0214]

Claims

1. A composition that inhibits the production of type 2 helper T cell (Th2)-mediated cytokines or alleviates allergic reactions, the composition comprising a Lactobacillus plantarum strain belonging to the genus Lactobacillus as an active ingredient.

2. A Lactobacillus plantarum GBCC_F0070 strain belonging to the genus Lactobacillus, with a deposit number of KCTC15051BP.

3. The strain according to claim 2, wherein, the strain comprises the 16S rRNA of SEQ ID NO:

1.

4. The strain according to claim 2, wherein, the strain comprises the recA gene sequence of SEQ ID NO:

2.

5. The strain according to claim 2, wherein, the strain comprises a mutation of a naturally occurring Lactobacillus plantarum strain.

6. The strain according to claim 2, wherein, the strain is a live bacterium, a dead bacterium, an attenuated bacterium, or a cytoplasmic fraction obtained by disrupting the strain.

7. The strain according to claim 2, wherein, the strain comprises the following characteristics: inhibiting Th2-mediated cytokine production; inhibiting the degranulation activity of mast cells; inhibiting the IgE level in serum; or inhibiting the MCPT-1 level in serum.

8. The strain according to claim 7, wherein, the Th2-mediated cytokine is any one or more selected from the group consisting of IL-4, IL-5, IL-6, IL-10, and IL-13.

9. A composition comprising a lysate, a culture broth, or an extract of the culture broth derived from the strain of claim 2.

10. The composition according to claim 9, wherein, the composition is used for preventing or treating IgE-mediated diseases or disorders.

11. The composition according to claim 9, wherein, the composition reduces the concentration of IgE in serum or reduces the binding of IgE to the surface receptor of mast cells.

12. The composition according to claim 9, wherein, the composition further comprises an additive selected from the group consisting of preservatives, dyes, emulsifiers, sweeteners, stabilizers, flavor enhancers, flavoring agents, and acidulants.

13. The composition according to claim 9, wherein, the composition is one or more dosage forms selected from the group consisting of solutions, emulsions, viscous mixtures, powders, granules, tablets, and capsules.

14. A health food composition for improving or preventing allergic diseases, comprising the Lactobacillus plantarum GBCC_F0070 strain deposited under the deposit number KCTC15051BP, a lysate, a culture broth, or a mixture thereof derived from the strain as an active ingredient.

15. The health food composition according to claim 14, wherein, the improvement of the allergic diseases includes: inhibiting Th2-mediated cytokine production; inhibiting the degranulation activity of mast cells; inhibiting the IgE level in serum; or inhibiting the MCPT-1 level in serum.

16. The strain according to claim 15, wherein, the Th2-mediated cytokine is any one or more selected from the group consisting of IL-4, IL-5, IL-6, IL-10, and IL-13.

17. The health food composition according to claim 14, wherein, the improvement of the allergic disease includes alleviating immune hypersensitivity or improving skin conditions caused by immune hypersensitivity.

18. The health food composition according to claim 14, wherein, the allergic disease is selected from the group consisting of edema, allergic reaction, allergic rhinitis, asthma, allergic conjunctivitis, allergic dermatitis, atopic dermatitis, contact dermatitis, urticaria, chronic spontaneous urticaria, pruritus, anaphylactic shock, insect allergy, food allergy, drug allergy, and respiratory allergy.

19. The health food composition according to claim 14, wherein, the allergic disease includes allergic reactions caused by indoor dust, fungi, mites, or animal hair, skin, or excrement.

20. A pharmaceutical composition for treating or preventing allergic diseases, which comprises Lactobacillus plantarum GBCC_F0070 strain deposited under the accession number KCTC15051BP, a lysate, a culture broth, or a mixture thereof derived from the strain as an active ingredient.

21. The pharmaceutical composition according to claim 20, wherein, the composition has mast cell degranulation inhibitory activity.

22. The pharmaceutical composition according to claim 20, wherein, the composition inhibits the expression of i) IgE or ii) the level of MCPT-1 in serum.

23. The pharmaceutical composition according to claim 20, wherein, the allergic disease is selected from the group consisting of edema, allergic reaction, allergic rhinitis, asthma, allergic conjunctivitis, allergic dermatitis, atopic dermatitis, contact dermatitis, urticaria, chronic spontaneous urticaria, pruritus, insect allergy, food allergy, drug allergy, and respiratory allergy.

24. The pharmaceutical composition according to claim 20, wherein, the allergic disease includes allergic reactions caused by indoor dust, fungi, mites, or animal hair, skin, or excrement.

25. The pharmaceutical composition according to claim 20, wherein, the composition can be administered orally or parenterally.

26. The pharmaceutical composition according to claim 25, wherein, the oral or parenteral administration is once a month, once every two weeks, once a week, once a day, twice a day, or three times a day.

27. The pharmaceutical composition according to claim 26, wherein, the parenteral administration is subcutaneous, intradermal, intravenous, intraarterial, intramuscular, intraperitoneal, intrathecal, intra-articular, intrasynovial, intrasternal, intralesional, or rectal administration.

28. The pharmaceutical composition according to claim 20, wherein, the composition is formulated for oral, rectal, intravenous, intravascular, intratumoral, subcutaneous, intradermal, or intraperitoneal delivery.

29. A cosmetic composition for improving or preventing skin allergic diseases, which comprises Lactobacillus plantarum GBCC_F0070 strain deposited under the accession number KCTC15051BP, a lysate, a culture broth, or a mixture thereof derived from the strain.

30. A method for preventing or treating allergic diseases, which comprises: An effective amount of a composition is administered to an individual in need thereof, the composition comprising Lactobacillus plantarum strain GBCC_F0070 deposited under accession number KCTC15051BP, a culture of the strain, a lysate of the strain, or a mixture thereof.

31. Use of a composition comprising Lactobacillus plantarum strain GBCC_F0070 deposited under accession number KCTC15051BP, a culture of the strain, a lysate of the strain, or a mixture thereof, in the manufacture of a pharmaceutical preparation for preventing or treating allergic diseases.

Images