KR102542226B1 - Composition for preventing or treating inflammatory bowel diseases - Google Patents

Abstract

The present invention relates to a composition for preventing or treating inflammatory bowel disease, and more particularly, to a composition for Tetragenococcus halophilus. two or more strains selected from the group consisting of halophilus ), Eubacterium rectale and Lactobacillus animalis ; It provides a pharmaceutical composition and a health food composition for preventing or treating inflammatory bowel disease, comprising a culture of the strains, a concentrate of the culture, an extract of the culture, or a lysate thereof as an active ingredient.
The composition can prevent, improve, or treat inflammatory bowel disease more effectively than a single strain by containing two or more strains having anti-inflammatory activity as active ingredients and regulating immune cells related to inflammatory bowel disease.

Description

Composition for preventing or treating inflammatory bowel disease {COMPOSITION FOR PREVENTING OR TREATING INFLAMMATORY BOWEL DISEASES}

The present invention relates to a composition for preventing or treating inflammatory bowel disease, and more particularly, to a composition for preventing or treating inflammatory bowel disease comprising two or more strains as active ingredients.

Inflammatory bowel disease (IBD) is a disease in which abnormal chronic inflammation in the intestinal tract repeatedly improves and recurs. Ulcerative colitis and Crohn's disease are typical, and the pathogenesis is still unclear. not revealed Inflammation diagnosis is made by synthesizing clinical symptoms, endoscopic and histopathological findings, blood test findings, and radiological examination findings. Inflammatory bowel disease is a chronic disease in which remission and exacerbation of symptoms are repeated, and its treatment is mainly aimed at controlling symptoms, preventing complications, and improving quality of life rather than curing the disease.

Crohn's disease is an inflammatory disease of unknown cause that occurs in the digestive tract from the mouth to the anus. Ulcerative colitis, the exact cause of which has not yet been identified, is characterized by chronic inflammation in the large intestine and repeated improvement and exacerbation due to a complex action of genetic predisposition, intestinal microbes and immunological abnormalities in the wall of the large intestine. there is

There is a need to develop a new therapeutic agent capable of effectively treating inflammatory bowel disease through the identification of mechanisms for the occurrence and control of inflammatory bowel disease and the regulation of related factors.

Republic of Korea Patent Publication No. 10-2011-0125667 (published on November 21, 2011)

An object of the present invention is to provide pharmaceutical compositions and health food compositions containing strains effective for preventing, improving or treating inflammatory bowel disease.

In order to achieve the above object, the present invention is Tetragenococcus halophilus ( Tetragenococcus halophilus ), Eubacterium rectale ( Eubacterium rectale ) and Lactobacillus animalis ( Lactobacillus animalis ) Two or more strains selected from the group consisting of; Provided is a pharmaceutical composition for preventing or treating inflammatory bowel disease comprising a culture of the strains, a concentrate of the culture, an extract of the culture, or a lysate thereof as an active ingredient.

In addition, the present invention is Tetragenococcus halophilus ( Tetragenococcus halophilus ), Eubacterium rectale ( Eubacterium rectale ) and Lactobacillus animalis ( Lactobacillus animalis ) Two or more strains selected from the group consisting of; It provides a health food composition for preventing or improving inflammatory bowel disease, comprising a culture of the strains, a concentrate of the culture, an extract of the culture, or a lysate thereof as an active ingredient.

The composition according to the present invention can prevent, improve, or treat inflammatory bowel disease more effectively than a single strain by including two or more strains having anti-inflammatory activity as active ingredients and regulating immune cells related to inflammatory bowel disease.

In addition, by combining the composition with an existing therapeutic agent, its therapeutic effect can be further enhanced.

1 is a graph showing changes in body weight of a mouse model for dextran sulfate sodium salt (DSS)-induced enteritis according to an embodiment of the present invention.
Figure 2 is a comparison of the colon length of the DSS-induced enteritis mouse model according to an embodiment of the present invention.
Figure 3 shows changes in the intestinal tissue of the DSS-induced enteritis mouse model according to an embodiment of the present invention.
Figure 4 shows the change in CD11b+ cell frequency in the DSS-induced enteritis mouse model according to another embodiment of the present invention. (PBL: Peripheral blood lymphocytes, peripheral blood lymphocytes / spleen: spleen)
Figure 5 shows the change in Ly6G+ cell frequency in the DSS-induced enteritis mouse model according to another embodiment of the present invention.
Figure 6 shows the change in the frequency of CD8 + NK1.1 + cells in the DSS-induced enteritis mouse model according to another embodiment of the present invention.
7 shows changes in the frequency of CD11b+Ly6G+ cells in a DSS-induced enteritis mouse model according to another embodiment of the present invention.
FIG. 8 shows changes in the cell frequency of FIGS. 6 and 7 in the spleen of the mouse model.
9 shows changes in the frequency of CD80+ cells in a DSS-induced enteritis mouse model according to another embodiment of the present invention. (pMQ: peritoneal macrophages, peritoneal macrophages)

Hereinafter, the present invention will be described in detail.

In the process of inducing enteritis with DSS (Dextran Sulfate Sodium Salt), the present inventors found that the symptoms of enteritis in mice were alleviated when various strains were orally administered, and the strain or a combination thereof was used to treat inflammatory bowel disease such as Crohn's disease or ulcerative colitis. By confirming that it can be utilized as a therapeutic agent or therapeutic adjuvant for a disease, the present invention was completed.

The present invention is Tetragenococcus halophilus ( Tetragenococcus halophilus ), Eubacterium rectale ( Eubacterium rectale ) and Lactobacillus animalis ( Lactobacillus animalis ) Two or more strains selected from the group consisting of; Provided is a pharmaceutical composition for preventing or treating inflammatory bowel disease comprising a culture of the strains, a concentrate of the culture, an extract of the culture, or a lysate thereof as an active ingredient.

In the present specification, "Tetragenococcus halophilus ( Tetragenococcus halophilus ; T. halophilus )" is a halophilic lactic acid bacterium (LAB) found in the fermentation process of soy sauce, soybean paste, fish sauce, and salted anchovies, also known as Pediococcus halophilus . do.

Tetragenococcus halophilus is a Gram-positive cocci of 0.6 to 1 μm, and has the characteristics of not being motile and not forming spores. The Tetragenococcus halophilus is a vegetable lactic acid bacterium, and compared to animal lactic acid bacteria that live in animal foods such as milk and yogurt, it can withstand acidic digestive juices in the stomach or small intestine, and has effects such as promoting intestinal activity, enhancing immunity, and removing harmful bacteria in the intestine. It is similar to animal lactic acid bacteria, but its vitality is more than 400 times stronger. In addition, the ability of vegetable lactic acid bacteria to remove aflatoxin, a toxin from mold, is eight times higher than that of animal lactic acid bacteria.

In the present specification, "Eubacterium Rectale ( Eubacterium rectale ; E. rectale )" is a microorganism that is a species of Eubacterium and is one of the most abundant bacteria in human excrement, and was found in association with rectal ulcers. The Eubacterium is a genus of Gram-positive anaerobic bacilli. As a genus, it has been isolated as a normal flora from feces, rumen, and periodontal tissue. The optimum growth temperature of the Eubacterium is 37° C., and the optimum pH is 7.0.

In the present specification, " Lactobacillus animalis ; L. animalis " is one of the most frequently occurring lactic acid bacteria in the manufacturing process of kimchi, a traditional Korean food. Lactobacillus is a gram-positive facultative anaerobe or microaerophilic bacterium that obtains energy by fermenting sugars and produces a large amount of lactic acid. represents polymorphism. The size of Lactobacillus genus is 0.5~0.9×1~11μm, and it shows various forms from short bacillus to enter bacilli.

In the present invention, the composition may include two or more strains selected from the group consisting of Tetragenococcus halophilus, Eubacterium rectale and Lactobacillus animalis, preferably Tetragenococcus halophilus, It may include both Eubacterium rectale and Lactobacillus animalis, but is not limited thereto.

More preferably, the composition has a bacterial count ratio of 1: (2 to 5): (0.2 to 0.6), preferably 1: ( 3 to 4): (0.3 to 0.5) may be included in the ratio of the number of bacteria, but is not limited thereto.

By including two or more of the above strains, the composition may further enhance improvement or treatment effects on inflammatory bowel disease.

In addition to the strains, the composition may include cultures of the strains, concentrates of the cultures, extracts of the cultures, or lysates thereof.

The culture may include a culture medium itself cultured in a medium, a filtrate obtained by removing the strain by filtering or centrifuging the culture medium, and a processed product derived from the culture medium itself, such as a concentrate or extract thereof.

The lysate may include a lysate obtained by disrupting the strain and a supernatant obtained by centrifuging the lysate. In addition, the composition may include a mixture of live, ie active strains and killed strains, but is not limited thereto.

The composition may further comprise one or more suitable prebiotic compounds.

In the present specification, a "prebiotic compound" is a non-digestible component that increases the growth of specific microorganisms in the gastrointestinal tract, and can become a nutrient source for probiotics to activate probiotics or increase the number of strains. there is. The prebiotic compound is resistant to hydrolysis and digestion and absorption by enzymes such as mammalian stomach acid, is fermented by the action of the probiotics, and can be a substance that selectively stimulates the growth and activity of the probiotics.

The prebiotic compounds are generally oligo-, polysaccharides, or non-digestible carbohydrates such as sugar alcohols, such as, for example, fructo-oligosaccharides (FOS), isomalto-oligosaccharides. (Isomalto-Oligosaccharide; IMO), Xylo-Oligosaccharide (XOS), Chito-Oligosaccharide (COS), Pectin, Inulin, Beta-Glucan (β -glucan), polydextrose, D-tagatose, and at least one selected from the group consisting of acacia fibers.

The amount and activity characteristics of the prebiotic compound may vary depending on the type, type, combination, etc. of the probiotics. For example, the composition may include the prebiotic compound in an amount of about 1 to about 30% by weight, preferably 5 to 20% by weight, based on 100% by weight of the total composition, and the prebiotic compound The activity of the strains can be changed by

In the present invention, the composition can regulate the expression of one or more immune cells whose phenotype is selected from the group consisting of CD11b+, Ly6G+, CD8+NK1.1+, CD11b+Ly6G+ and CD80+, preferably the above immune cells. can be reduced or inhibited, or the expression of the CD11b+, Ly6G+, CD8+NK1.1+, CD11b+Ly6G+ or CD80+ can be reduced.

According to one embodiment of the present invention, it can be confirmed that the strains have anti-inflammatory activity by reducing the expression of immune cells and their activating molecules related to inflammation.

In the present invention, the inflammatory bowel disease may be at least one selected from the group consisting of ulcerative colitis, Crohn's disease, Behçet's disease enteritis, infectious enteritis, radiation-induced enteritis, ischemic bowel disease, and irritable bowel syndrome, but is limited thereto It is not.

The infectious enteritis may include bacterial, viral, amoebic, or tuberculous enteritis, but is not limited thereto.

The pharmaceutical composition according to the present invention can be prepared according to conventional methods in the pharmaceutical field. The pharmaceutical composition may be formulated with an appropriate pharmaceutically acceptable carrier according to the formulation and, if necessary, may further contain excipients, diluents, dispersants, emulsifiers, buffers, stabilizers, binders, disintegrants, solvents, and the like. there is. The appropriate carrier and the like do not inhibit the activity and characteristics of the strains according to the present invention, and may be selected differently depending on the dosage form and formulation.

The pharmaceutical composition may be applied in any dosage form, and more specifically, it may be formulated and used in parenteral dosage forms such as oral dosage forms, external preparations, suppositories and sterile injection solutions according to conventional methods.

Among the oral dosage forms, the solid dosage form is in the form of tablets, pills, powders, granules, gels, capsules, etc., and includes at least one excipient, for example, starch, calcium carbonate, sucrose, lactose, sorbitol, mannitol, cellulose, It can be prepared by mixing gelatin, etc., and lubricants such as magnesium stearate and talc may be included in addition to simple excipients. In addition, in the case of a capsule formulation, a liquid carrier such as fatty oil may be further included in addition to the above-mentioned materials.

Among the oral formulations, liquid formulations include suspensions, solutions for internal use, emulsions, syrups, etc. In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, aromatics, and preservatives may be included. there is.

The parenteral formulation may include a sterilized aqueous solution, a non-aqueous solvent, a suspension, an emulsion, a lyophilized formulation, and a suppository. Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used as non-aqueous solvents and suspending agents. As a base for the suppository, witepsol, macrogol, Tween 61, cacao butter, laurin fat, glycerogeratin, and the like may be used. It is not limited thereto, and all suitable agents known in the art may be used.

In addition, calcium or vitamins may be further added to the pharmaceutical composition according to the present invention to enhance therapeutic efficacy.

The pharmaceutical composition according to the present invention can be administered in a pharmaceutically effective amount.

In the present specification, "pharmaceutically effective amount" means an amount that is sufficient to treat a disease with a reasonable benefit/risk ratio applicable to medical treatment and does not cause side effects.

The effective dosage level of the pharmaceutical composition depends on the purpose of use, the patient's age, sex, weight and health condition, disease type, severity, drug activity, drug sensitivity, administration method, administration time, administration route and excretion rate, treatment Duration, combination, or factors including drugs used concurrently and other factors well known in the medical arts may be determined differently. For example, a suitable daily dose of bacteria for an adult human can be about 1 x 10 ³ to about 1 x 10 ¹¹ CFU (colony forming units), eg about 1 x 10 ⁶ to about 1 x 10 ¹⁰ CFU, in another example, about 1 x 10 ⁷ to about 1 x 10 ⁸ CFU.

The pharmaceutical composition may be administered individually or in combination with other therapeutic agents, or may be administered sequentially or simultaneously with conventional therapeutic agents. In addition, single or multiple administrations may be possible. It is important to administer the amount that can obtain the maximum effect with the minimum amount without side effects in consideration of all the above factors.

The pharmaceutical composition may be administered to any animal that may develop inflammatory bowel disease, and the animal may include, for example, humans and primates as well as livestock such as cattle, pigs, horses, and dogs.

The pharmaceutical composition may be administered by an appropriate administration route according to the formulation form, and may be administered through various oral or parenteral routes as long as it can reach the target tissue. The method of administration is not particularly limited, and may be administered by conventional methods such as oral, rectal or intravenous, intramuscular, skin application, intraventricular inhalation, intrauterine dural or intracerebroventricular injection. there is.

The pharmaceutical composition may be used alone for the prevention or treatment of inflammatory bowel disease, or may be used in combination with surgery or other drug treatment, and compounds commonly used in the treatment of inflammatory bowel disease, such as antibiotics, anti-inflammatory agents, It can be particularly effective when used in combination with such additional therapeutic agents as anesthetics, analgesics, and the like.

The present invention is Tetragenococcus halophilus ( Tetragenococcus halophilus ), Eubacterium rectale ( Eubacterium rectale ) and Lactobacillus animalis ( Lactobacillus animalis ) Two or more strains selected from the group consisting of; It provides a health food composition for preventing or improving inflammatory bowel disease, comprising a culture of the strains, a concentrate of the culture, an extract of the culture, or a lysate thereof as an active ingredient.

Corresponding features may be substituted in the foregoing.

For the purpose of preventing or improving inflammatory bowel disease, the health food according to the present invention may be prepared in powder, granule, tablet, capsule, syrup or beverage form. There is no limit to the form that the health food can take, and it can be formulated in the same way as the pharmaceutical composition and used as a functional food or added to various foods.

The health food may include all foods in a conventional sense. For example, beverages and various drinks, fruits and their processed foods (canned fruit, jam, etc.), fish, meat and their processed foods (ham, bacon, etc.), breads and noodles, cookies and snacks, dairy products (butter, cheese, etc.) ), etc., and may include all functional foods in a conventional sense. In addition, food used as feed for animals may also be included.

The health food composition may be prepared by further including food chemically acceptable food additives (food additives) commonly used in the art and other suitable auxiliary ingredients. The suitability as a food additive can be determined according to the standards and standards for the item in accordance with the general rules of the Food Additive Code and general test methods approved by the Ministry of Food and Drug Safety, unless otherwise specified. Examples of the items listed in the 'Food Additive Code' include, for example, chemical compounds such as ketones, glycine, calcium citrate, nicotinic acid, and cinnamic acid; natural additives such as persimmon pigment, licorice extract, crystalline cellulose, kaoliang pigment, and guar gum; mixed preparations such as sodium L-glutamate preparations, noodle-added alkali preparations, preservative preparations, and tar color preparations; and the like.

The other auxiliary ingredients include, for example, flavoring agents, natural carbohydrates, sweeteners, vitamins, electrolytes, colorants, pectic acid, alginic acid, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents, etc. may additionally contain. In particular, as the natural carbohydrate, monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol may be used. As the sweetener, natural sweeteners such as thaumatin and stevia extract or synthetic sweeteners such as saccharin and aspartame may be used.

An effective dose of the strains contained in the health food may be appropriately adjusted according to the purpose of use, such as prevention or improvement of inflammatory bowel disease.

The health food composition uses food as a raw material, has no side effects that can occur when taking general medicines for a long time, has the advantage of providing nutritional benefits, and is excellent in portability, for preventing or improving inflammatory bowel disease. It can be taken as a supplement.

In addition, the present invention is Tetragenococcus halophilus ( Tetragenococcus halophilus ), Eubacterium rectale ( Eubacterium rectale ) and Lactobacillus animalis ( Lactobacillus animalis ) Two or more strains selected from the group consisting of; Provided is a composition for enhancing the therapeutic effect of a therapeutic agent for inflammatory bowel disease, comprising a culture of the strains, a concentrate of the culture, an extract of the culture, or a lysate thereof as an active ingredient.

Corresponding features may be substituted in the foregoing.

Hereinafter, examples will be described in detail to aid understanding of the present invention. However, the following examples are merely illustrative of the contents of the present invention, but the scope of the present invention is not limited to the following examples. The embodiments of the present invention are provided to more completely explain the present invention to those skilled in the art.

< Example 1> Preparing strains

Strains used in the following experimental examples were distributed from KCTC and ATCC.

- Tetragenococcus Halophilus ( Tetragenococcus halophilus ): KCTC #3720

- Lactobacillus animalis ( Lactobacillus animalis ): KCTC # 3501

- Eubacterium rectale ( Eubacterium rectale ): ATCC® 33656™, KCTC #5835

< Example 2> Confirmation of the treatment effect of inflammatory bowel disease

After administering the bacterial strain according to Example 1 to a mouse model of DSS (Dextran Sulfate Sodium Salt)-induced enteritis, the change in body weight and immune cell phenotype were confirmed to suppress the aggravation of symptoms of DSS-induced enteritis and confirm the immune mechanism related thereto. did

Comparison between a group in which enteritis was induced by oral administration of DSS for 1 week and additional drinking water for 3 days to mice, and a group in which DSS and bacterial strains were administered simultaneously for 1 week and bacterial strains were administered for 3 additional days. did

The bacterial strain administration group consisted of a single administration group, a two-type bacteria complex administration group, and a three-type bacteria complex administration group. The doses were 1 x 10 ⁸ cfu of Eubacterium Rectale, 3.8 x 10 ⁸ cfu of Tetragenococcus Halophilus, and 3.9 x 10 ⁷ cfu of Lactobacillus Animalis. For each strain, an administration ratio of 1:3.8:0.39 was applied. A normal control was administered drinking water. After that, the mice were used in the following experiments.

1) Confirmation of body weight change in the DSS-induced enteritis mouse model and body weight change in the group administered with the strain

As a result of confirming the change in body weight of the DSS-administered mice, as shown in FIG. 1, the body weight of the DSS-administered mice decreased to a statistically significant level (p=0.001), but in the control mice and the group administered with DSS and bacterial strains There was no significant decrease in body weight. After 10 days of administration, there was a statistically significant increase in body weight compared to the DSS administration group.

2) Comparison of colon length between the DSS-induced enteritis mouse model and the group administered with the strain

As a result of comparing the colon length of DSS-administered mice, as shown in FIG. 2, the length of the colon decreased in the DSS-induced enteritis mouse model, but when the bacterial strain was administered, the colon length increased statistically significantly and was recovered. appear.

3) Comparison of the histology of intestinal tissue in the DSS-induced enteritis mouse model and the group administered with the bacterial strain

As a result of comparing the intestinal tissue morphology of the DSS-administered mice, as shown in FIG. 3, it was confirmed that the intestinal tissue morphology of the group simultaneously administered with the three strains was maintained at the most similar level to that of normal mice compared to the DSS-induced enteritis mouse model. there was.

< Example 3> Confirmation of inflammatory bowel disease immunomodulatory effect

1) CD11 is involved in adhesion and inflammation between cells such as monocytes, macrophages, natural killer (NK) cells and granulocytes. Referring to FIG. 4, it was found that the frequency of CD11b+ cells decreased in the group in which three strains were simultaneously administered among the DSS-induced enteritis mouse models.

2) Ly6G (Lymphocyte antigen 6 complex locus G6D) is a marker for monocytes, granulocytes and neutrophils. Referring to FIG. 5, Ly6G+ granulocytes increased in the DSS-induced enteritis mouse model, and the frequency of Ly6G+ cells was most effectively reduced when two or three bacterial strains were administered.

3) CD8+NK1.1+ cells play a pathogenic role and are known as the main target for disease control. 6 and 8, the frequency of CD8+NK1.1+ cells increased in the DSS-induced enteritis mouse model, but the frequency of CD8+NK1.1+ cells decreased in the group administered with two or three strains. In particular, a significant decrease was shown in the group administered with three strains.

4) CD11b+Ly6G+ cells are used as a neutrophil marker, and an increase in neutrophils is a characteristic of an inflammatory response. 7 and 8, the frequency of CD11b+Ly6G+ cells increased in the DSS-induced enteritis mouse model, but the frequency of these cells decreased in the group administered with two or three strains, particularly The most significant decrease was shown in the three strains administered group.

5) CD80+ cells in the colonic mucosa of Crohn's disease and ulcerative colitis are significantly increased compared to the control group. It is known that the frequency of CD80+ cells decreases when the DSS-induced enteritis mouse model improves. Referring to FIG. 9 , the frequency of CD80+ cells was significantly decreased among intraperitoneal cells of the group in which the three strains were administered to the DSS-induced enteritis mouse model.

Having described specific parts of the present invention in detail above, it is clear to those skilled in the art that these specific descriptions are only preferred embodiments, and the scope of the present invention is not limited thereby. do. That is, the substantial scope of the present invention is defined by the appended claims and their equivalents.

Claims (8)

a strain mixture consisting of Tetragenococcus halophilus , Eubacterium rectale and Lactobacillus animalis ; A pharmaceutical composition for preventing or treating inflammatory bowel disease comprising a culture of the strain mixture, a concentrate of the culture, an extract of the culture, or a lysate thereof as an active ingredient. delete According to claim 1,
The composition,
A pharmaceutical composition comprising Eubacterium rectale, Tetragenococcus halophilus, and Lactobacillus animalis in a bacterial count ratio of 1: (2 to 5): (0.2 to 0.6). According to claim 1,
The composition,
A pharmaceutical composition characterized in that the phenotype decreases one or more immune cells selected from the group consisting of CD11b+, Ly6G+, CD8+NK1.1+, CD11b+Ly6G+ and CD80+. According to claim 1,
The inflammatory bowel disease,
Ulcerative colitis, Crohn's disease, Behcet's disease enteritis, infectious enteritis, radiation enteritis, ischemic bowel disease, characterized in that at least one selected from the group consisting of irritable bowel syndrome, a pharmaceutical composition. a strain mixture consisting of Tetragenococcus halophilus , Eubacterium rectale and Lactobacillus animalis ; A health food composition for preventing or improving inflammatory bowel disease comprising a culture of the strain mixture, a concentrate of the culture, an extract of the culture, or a lysate thereof as an active ingredient. delete According to claim 6,
The inflammatory bowel disease,
Ulcerative colitis, Crohn's disease, Behcet's disease enteritis, infectious enteritis, radiation enteritis, ischemic bowel disease, characterized in that at least one selected from the group consisting of, and irritable bowel syndrome, health food composition.

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