Lactic acid bacteria contain components

Abstract

To provide novel effects and uses of lactic acid bacteria.SOLUTION: The present invention provides a composition for improving alveolar bone metabolism containing Lactiplantibacillus plantarum S25 strain (accession number: NITE ABP-03571).SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present disclosure relates to a composition containing lactic acid bacteria and uses thereof, etc. The contents of all documents described in this specification are incorporated herein by reference. [Background technology]

[0002] Lactic acid bacteria fermentation products have been investigated for their various effects, such as improving the intestinal barrier (Patent Document 1) and improving skin condition (Patent Document 2), and useful lactic acid bacteria are being sought. [Prior art documents] [Patent documents]

[0003] [Patent Document 1] Japanese Patent Application Publication No. 2019-011315 [Patent Document 2] Japanese Patent Application Publication No. 2019-011316 Summary of the Invention [Problem to be solved by the invention]

[0004] The present inventors have explored new effects and uses of lactic acid bacteria.

[0005] The present inventors have identified Lactiprancibacillus Genus lactic acid bacteria (Lactiplantibacillus s) We found that ingesting the S25 strain (accession number: NITE ABP-03571) can suppress the production of inflammatory cytokines that activate osteoclasts in gingival tissue, and further improvements were made.

[0006] The present disclosure includes, for example, the subject matter described in the following sections: Section 1. Lactipranchibacillus Genus lactic acid bacteria (Lactiplantibacillus s)A composition for improving alveolar bone metabolism containing S25 strain (received number: NITE ABP-03571). Section 2. Item 1. The composition according to Item 1, which is used to inhibit osteoclasts in alveolar bone and / or to inhibit alveolar bone resorption. Section 3. Lactipranchibacillus Genus lactic acid bacteria (Lactiplantibacillus s) A composition for improving or maintaining periodontal tissues containing the S25 strain (accession number: NITE ABP-03571). Section 4. Item 4. The composition according to Item 3, wherein the periodontal tissue is alveolar bone or gingiva. Section 5. Item 5. The composition according to any one of Items 1 to 4, which is an oral composition. Section 6. Item 6. The composition according to any one of Items 1 to 5, which is a food or drink composition or a pharmaceutical composition. [Effects of the Invention]

[0007] Lactipranchibacillus Genus lactic acid bacteria (Lactiplantibacillus s) The present invention provides a composition for improving alveolar bone metabolism, which contains the S25 strain. Genus lactic acid bacteria (Lactiplantibacillus s) A composition for improving or maintaining periodontal tissues containing the S25 strain is provided. [Brief explanation of the drawings]

[0008] [Figure 1A] The results of measuring TNF-α gene expression in gingival tissues of mice orally fed a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (HFD+S25) are shown. * indicates p<0.05, and ** indicates p<0.01. [Figure 1B]The results of measuring COX-2 gene expression in gingival tissues of mice orally fed a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (HFD+S25) are shown. Note that + indicates p<0.1 and * indicates p<0.05. [Figure 1C] The results of measuring IL-8 gene expression in gingival tissues of mice orally fed a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (HFD+S25) are shown. * indicates p<0.05. [Figure 1D] This figure shows the results of measuring PAI-1 gene expression in gingival tissues of mice orally fed a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (HFD+S25). Note that + indicates p<0.1. [Figure 1E] The results of measuring MCP-1 gene expression in gingival tissues of mice orally fed a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (HFD+S25) are shown. Note that + indicates p<0.1 and * indicates p<0.05. [Figure 2A] The figures show the results of measuring the number of osteoclasts around the alveolar bone in mice orally fed a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (HFD+S25). * indicates p<0.05, and ** indicates p<0.01. [Figure 2B] The results of TRAP staining around the alveolar bone of mice orally fed a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (S25) are shown. DETAILED DESCRIPTION OF THE INVENTION

[0009] Each embodiment included in the present disclosure will be described in further detail below. The composition encompassed by the present disclosure is an oral composition containing lactic acid bacteria. In this specification, the composition may be referred to as the "composition of the present disclosure."

[0010] Lactic acid bacteria encompassed by the present disclosure include Lactiprancibacillus Genus lactic acid bacteria (Lactiplantibacillus s) S25 strain (received number: NITE ABP-03571). Lactipranchibacillus Genus lactic acid bacteria The S25 strain was received on December 15, 2021, at the Patent Microorganism Deposit Center of the National Institute of Technology and Evaluation (2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, Japan) under the receipt number NITE ABP-03571.

[0011] The lactic acid bacteria of the present disclosure are plant-derived lactic acid bacteria isolated from suguki, a Kyoto pickle.

[0012] The content of lactic acid bacteria in the composition of the present disclosure is not particularly limited, and is, for example, preferably about 0.1 to 100% by mass, more preferably about 1 to 99% by mass, 2 to 90% by mass, or 5 to 50% by mass.

[0013] In the composition of the present disclosure, the lactic acid bacteria may be viable or dead, with dead bacteria being preferred.

[0014] In the composition of the present disclosure, the lactic acid bacteria may or may not have been subjected to, for example, crushing, heating, drying (freeze drying, vacuum drying, spray drying, etc.), freezing, lysis, extraction, etc.

[0015] The composition of the present disclosure contains the above-mentioned lactic acid bacteria and may further contain other ingredients. Examples of such other ingredients include pharmacologically or food hygienically acceptable bases, carriers, solvents, dispersants, emulsifiers, buffers, stabilizers, excipients, binders, disintegrants, lubricants, thickeners, antioxidants, preservatives, coating agents, colorants, etc. These ingredients may be used alone or in combination of two or more.

[0016] The composition of the present invention is not particularly limited, but may be, for example, an oral composition.

[0017] The composition of the present invention may be a pharmaceutical composition or a food or drink composition. In the case of a food or drink composition, it may be, for example, a processed food, a beverage, a health food (a food with nutrient function claims, a food for specified health uses, etc.), a supplement, a food for the sick (a hospital food, a food for sick people, a nursing care food, etc.), etc.

[0018] The composition of the present disclosure can be prepared by a conventional method by combining the above-mentioned lactic acid bacteria with other ingredients as needed.

[0019] The composition of the present disclosure may have the effect of improving alveolar bone metabolism, inhibiting osteoclasts in alveolar bone, inhibiting alveolar bone resorption, reducing the number of osteoclasts, suppressing the expression of inflammatory cytokines (e.g., TNF-α (tumor necrosis factor-α), IL-8 (interleukin-8), MCP-1 (monocyte chemoattractant protein-1), etc.), suppressing the expression of adipocytokines (e.g., PAI-1 (plasminogen activator inhibitor-1), etc.), and / or suppressing the expression of COX-2 (cyclooxygenase-2). For this reason, the composition of the present disclosure is preferably used for improving alveolar bone metabolism, inhibiting osteoclasts in alveolar bone, inhibiting alveolar bone resorption, reducing the number of osteoclasts, inhibiting the expression of inflammatory cytokines (e.g., TNF-α, IL-8, MCP-1, etc.), inhibiting the expression of adipocytokines (e.g., PAI-1, etc.), and / or inhibiting the expression of COX-2. As used herein, "alveolar bone metabolism" refers to remodeling of the alveolar bone, in which bone formation by osteoblasts and bone resorption by osteoclasts are repeated in the alveolar bone. As used herein, "improving alveolar bone metabolism" refers to restoring bone metabolism in the alveolar bone, which is biased toward bone resorption, to a normal balance. As used herein, "suppressing osteoclasts in the alveolar bone" refers to suppressing differentiation into osteoclasts in the alveolar bone or suppressing the increase in osteoclasts in the alveolar bone.

[0020] Without wishing to be bound by theory, oral ingestion of the composition of the present disclosure can suppress the expression of the above-mentioned inflammatory cytokines and adipocytokines, particularly in the gums, thereby improving alveolar bone metabolism, suppressing osteoclasts in the alveolar bone, suppressing alveolar bone resorption (bone resorption in the alveolar bone), or reducing the number of osteoclasts in the alveolar bone.

[0021] Excessive production of inflammatory cytokines (e.g., TNF-α, IL-8, MCP-1, etc.) activates gingival fibroblasts and osteoclasts. Activated gingival fibroblasts produce collagenase, destroying gingival tissue, and activated osteoclasts promote alveolar bone resorption. These can cause gingival recession (sagging gums), making it impossible to maintain healthy periodontal tissues (e.g., alveolar bone, gingiva, etc.). For this reason, suppressing the expression of inflammatory cytokines is important.

[0022] PAI-I is a type of cytokine produced by mast cells and is known to be involved in the progression of thrombosis. There are also reports that there is a correlation between the amount of PAI-1 in the blood and periodontal condition, and it is thought that reduced blood flow to periodontal tissues may accelerate the progression of periodontal disease. For this reason, it is important to suppress the expression of PAI-1.

[0023] Furthermore, although not a cytokine, the composition of the present disclosure can also suppress the expression of COX-2, for example. COX-2 is induced by inflammatory cytokines and produces prostaglandins (PGs: lipid-based bioactive substances), which are thought to be involved in the cause of periodontal disease. PGs (especially PGE2 among PGs) activate gingival fibroblasts and osteoclasts, which can lead to the inability to maintain healthy periodontal tissues (e.g., alveolar bone, gingiva, etc.). For this reason, suppressing COX-2 expression is important.

[0024] From the above, the composition of the present disclosure can also be used for improving or maintaining periodontal tissues (for example, alveolar bone, gums, etc.).

[0025] Subjects that may take the compositions of the present disclosure include, for example, humans and non-human mammals (for example, rats, mice, rabbits, cows, pigs, dogs, cats, sheep, monkeys, etc.). Examples of humans include those with increased bone resorption in the alveolar bone, such as those whose bone metabolism is biased toward bone resorption due to aging or menopause; those with reduced alveolar bone; those with gingival inflammation (e.g., bleeding, redness, gingival swelling, etc.); those with gingival recession and the appearance of periodontal pockets; those with gingivitis; those with periodontitis; periodontal disease patients; those who grind or clench their teeth; those with partial tooth loss that places stress on surrounding teeth; those with toxins or inflammation caused by oral bacterial flora; and those with calcium or vitamin D deficiency. Furthermore, even if no significant abnormalities are observed in alveolar bone metabolism, the composition can also be used prophylactically to maintain normal alveolar bone metabolism. Furthermore, even if no significant abnormalities are observed in periodontal tissues (e.g., alveolar bone, gingiva, etc.), the composition can also be used prophylactically to maintain periodontal tissues (e.g., alveolar bone, gingiva, etc.).

[0026] The amount of lactic acid bacteria to be ingested is not particularly limited, and is preferably 1 to 100 billion per day (particularly per day for adults), and more preferably 1 to 10 billion per day. The amount of lactic acid bacteria contained in the composition of the present disclosure can also be determined based on the daily intake of lactic acid bacteria. 11 ~1.0~10 13 Therefore, for example, by using this value as a reference, the composition of the present disclosure can be prepared by adjusting the amount of killed lactic acid bacteria powder to contain the above-mentioned daily intake amount of lactic acid bacteria. Alternatively, the amount of lactic acid bacteria contained in the composition can be set to, for example, 1 / 2 or 1 / 3 of the above-mentioned daily intake amount of lactic acid bacteria, and the composition can be ingested several times a day (2 or 3 times).

[0027] It should be noted that in this specification, the term "comprising" includes "consisting essentially of" and "consisting of." Furthermore, the present disclosure encompasses all arbitrary combinations of the constituent elements described in this specification.

[0028] Furthermore, the various characteristics (properties, structures, functions, etc.) described in each embodiment of the present disclosure above may be combined in any way to specify the subject matter encompassed by the present disclosure, i.e., the present disclosure encompasses all subject matter consisting of any combination of the combinable characteristics described herein. [Example]

[0029] Hereinafter, the embodiments of the present disclosure will be described more specifically with reference to examples, but the embodiments of the present disclosure are not limited to the following examples.

[0030] Animals used Three-week-old male C57BL / 6J (SPF) mice were purchased from Charles River Japan. The acclimation period was from the day of arrival to the day of group allocation. The quarantine period lasted until the 7th day, with the day of arrival being considered day 0. Feed was provided ad libitum except during fasting periods, using D12450J (normal feed (Feed 1), RESEARCH DIETS, INC., lot number: 20080303) placed on the cage lid during the quarantine and acclimation periods and the test period for Group A, and D12492 (high-fat diet (HFD; Feed 2), RESEARCH DIETS, INC., lot numbers: 20090202, 20110303) during the test period for Groups B and C. Drinking water was provided ad libitum throughout the test period using a polysulfone waterer (stainless steel tip).

[0031] Grouping Using the body weight measured on the final day of the quarantine and acclimation period as an index, the animals were assigned to groups using a stratified sequential randomization method according to the following table.

[0032] [Table 1]

[0033] The test substance was Lactipranchibacillus Genus lactic acid bacteria The S25 strain (received number: NITE ABP-03571) (killed bacteria) was used. The dose is shown in mg / animal in the above table, but the number of bacteria is 10 hundred million The mice were given 1000 pieces per day per animal. Distilled water containing the test substance was administered via a probe. Mice in the non-test substance group received distilled water via a probe. The administration period was 77 days. After the administration period, the mice were fasted overnight, anesthetized, and dissected to collect gingival tissue (left and right maxillary sides, right mandibular side). After cutting the tissue in a frozen state with a scalpel, total RNA was collected using a Total RNA Mini Kit (Bio-Rad) and reverse-transcribed to produce cDNA. The following gene expression changes were then monitored by real-time PCR. The results are shown in Figures 1A-1E, where the gene expression level in the normal diet group (Group A: control) is set to 1. ·TNF-α (tumor necrosis factor-α) COX-2 (cyclooxygenase-2) IL-8 (Interleukin-8) PAI-1 (plasminogen activator inhibitor-1) ·MCP-1 (monocyte chemoattractant protein-1, monocyte chemotactic factor)

[0034] As shown in Figures 1A-E, Lactipranchibacillus Genus lactic acid bacteria In the group administered with S25 strain, the expression levels of TNF-α, COX-2, IL-8, PAI-1, and MCP-1 in gingival tissue were confirmed to be reduced.

[0035] The number of osteoclasts was assessed using TRAP staining (Acid Phosphatase, Leukocyte (TRAP) Kit (Sigma 387A)). Slides were immersed in a fixative (4% formaldehyde) and left to stand at room temperature for 30 seconds, then washed with purified water preheated to 37°C. 0.5 ml of Fast GBC Base Solution and 0.5 ml of Sodium Nitrite Solution were placed in a tube, which was then mixed by inversion for 30 seconds and left to stand for 2 minutes. The staining solution was prepared in a beaker, transferred to a staining bottle, and heated to 37°C in a thermostatic bath. The slides were then immersed in the staining bottle and incubated at 37°C for 1 hour in the dark. The slides were then washed with purified water and then washed with hematoxylin. The specimens were then immersed in the solution for 2 minutes for counterstaining. They were then rinsed with pure water for several minutes, air-dried, and mounted in glycerol gelatin (Sigma, GG1). The specimens were then observed under a microscope, and photographs of the periodontal tissues were taken. The number of osteoclasts present near the alveolar bone around the teeth was counted. The number of osteoclasts is shown in Figure 2A, and the results of TRAP staining are shown in Figure 2B.

[0036] As shown in Figure 2A and B, the intake of a high-fat diet was confirmed to increase the number of osteoclasts, and Lactipranchibacillus Genus lactic acid bacteria In the group administered with the S25 strain, this increase was suppressed.

Claims

1. A composition for improving alveolar bone metabolism, comprising Lactiplantibacillus S25 strain (accession number: NITE ABP-03571).

2. The composition according to claim 1, which is used to inhibit osteoclasts in alveolar bone and / or to inhibit alveolar bone resorption.

3. A composition for improving or maintaining periodontal tissues, comprising Lactiplantibacillus S25 strain (accession number: NITE ABP-03571).

4. The composition according to claim 3, wherein the periodontal tissue is alveolar bone or gingiva.

5. The composition according to any one of claims 1 to 4, which is an oral composition.

6. The composition according to any one of claims 1 to 5, which is a food or drink composition or a pharmaceutical composition.

Images