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 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 its uses, etc. The contents of all documents described in this specification are incorporated herein by reference.

Background Art

[0002] It has been studied that various effects are exerted by lactic acid bacteria fermentation products. For example, it is known that effects such as improvement of the intestinal barrier (Patent Document 1) or improvement of the skin (Patent Document 2) are exerted, and useful lactic acid bacteria are being searched for.

Prior Art Documents

Patent Documents

[0003]

Patent Document 1

Patent Document 2

Summary of the Invention

Problems to be Solved by the Invention

[0004] The present inventors searched for novel effects and uses of lactic acid bacteria.

[0005] The present inventors found that by ingesting Lactiplantibacillus Genus lactic acid bacteria (Lactiplantibacillu s) S25 strain (Accession No.: NITE ABP-03571), it is possible to suppress the production of inflammatory cytokines that activate osteoclasts, and further improvements were made.

[0006] The present disclosure includes, for example, the subject matter described in the following items. Item 1. Lactiplantibacillus Genus lactic acid bacteria (Lactiplantibacillu s)A composition for improving bone metabolism containing strain S25 (receipt number: NITE ABP-03571). Section 2. The composition according to item 1, for the purpose of inhibiting osteoclasts and / or inhibiting bone resorption. Section 3. The composition described in item 1 or 2, which is an oral composition. Section 4. A composition according to any one of items 1 to 3, which is a food or beverage composition or a pharmaceutical composition. [Effects of the Invention]

[0007] Lactiplantibacillus Genus lactic acid bacteria (Lactiplantibacillus s) A composition for improving bone metabolism containing the S25 strain is provided. [Brief explanation of the drawing]

[0008] [Figure 1A] The results of TNF-α gene expression measurements in gingival tissue of mice orally administered 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 COX-2 gene expression measurements in gingival tissue of mice orally administered a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (HFD+S25) are shown. + indicates p<0.1, and * indicates p<0.05. [Figure 1C] The results of IL-8 gene expression measurements in gingival tissue of mice orally administered 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] The results of PAI-1 gene expression measurements in gingival tissue of mice orally administered a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (HFD+S25) are shown. A "+" indicates p<0.1. [Figure 1E] The results of MCP-1 gene expression measurements in gingival tissue of mice orally administered a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (HFD+S25) are shown. + indicates p<0.1, and * indicates p<0.05. [Figure 2A] The results of measuring MCP-1 gene expression in adipose tissue of mice orally fed either a normal diet (Control) or a high-fat diet (HFD) are shown. ** indicates p<0.01. [Figure 2B] The results of PPARγ gene expression measurements in adipose tissue of mice orally administered 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 2C] The results of measuring adiponectin gene expression in adipose tissue of mice orally administered 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 3A] The results of measuring the number of osteoclasts around the alveolar bone in mice that orally ingested 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 3B] The results of TRAP staining around the alveolar bone of mice that were orally fed a normal diet (Control), a high-fat diet (HFD), or a high-fat diet and lactic acid bacteria (S25) are shown. [Modes for carrying out the invention]

[0009] The embodiments included in this disclosure will be described in further detail below. The compositions included in this disclosure are oral compositions containing lactic acid bacteria. In this specification, such compositions may be referred to as "the compositions of this disclosure."

[0010] The lactic acid bacteria included in the present disclosure are Lactiplantibacillus Genus lactic acid bacteria (Lactiplantibacillu s) strain S25 (accession number: NITE ABP - 03571). Lactiplantibacillus Genus lactic acid bacteria Strain S25 was received on December 15, 2021, at the Patent Microorganism Deposit Center of the National Institute of Technology and Evaluation (2 - 5 - 8 Kazusa Kamashima, Kisarazu City, Chiba Prefecture, Japan) with the accession number: NITE ABP - 03571.

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

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

[0013] In the composition of the present disclosure, the lactic acid bacteria may be live cells or dead cells. Among them, dead cells are preferable.

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

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

[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 beverage composition. If it is a food or beverage composition, it may be, for example, a processed food, a beverage, a health food (nutrient functional food, food for specified health uses, etc.), a supplement, or a food for the sick (hospital food, sick person's food, or nursing care food, etc.).

[0018] The compositions of this disclosure can be prepared by conventional methods by combining the lactic acid bacteria described above with other components as needed.

[0019] The compositions disclosed herein may exhibit effects such as improving bone metabolism, suppressing osteoclasts, suppressing 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.)), suppressing the expression of COX-2 (cyclooxygenase-2), enhancing the expression of PPARγ (Peroxisome Proliferator-Activated Receptor γ), and / or enhancing the expression of adiponectin. Therefore, the compositions of this disclosure are preferably used for improving bone metabolism, suppressing osteoclasts, inhibiting bone resorption, reducing the number of osteoclasts, suppressing the expression of inflammatory cytokines (e.g., TNF-α, IL-8, MCP-1, etc.), suppressing the expression of adipocytokines (e.g., PAI-1, etc.), suppressing the expression of COX-2, enhancing the expression of PPARγ, and / or enhancing the expression of adiponectin. Furthermore, the compositions of this disclosure can also be suitably used to balance bone metabolism; maintain strong bones; prevent poor posture and maintain appearance; maintain daily physical activity levels; prevent frailty and bedridden life, which are states of physical and mental decline associated with aging, and maintain quality of life. In this specification, "bone metabolism" means bone remodeling, which involves repeated bone formation by osteoblasts and bone resorption by osteoclasts. In this specification, "improvement of bone metabolism" means improving bone metabolism that is biased towards bone resorption to a normal balance. In this specification, "osteoclast suppression" means suppressing differentiation into osteoclasts or suppressing the increase of osteoclasts.

[0020] While we do not wish to be bound by theory, ingesting the composition of this disclosure may suppress the expression of inflammatory cytokines and adipocytokines, as described above, thereby improving bone metabolism, suppressing osteoclasts, inhibiting bone resorption, or reducing the number of osteoclasts.

[0021] When inflammatory cytokines (e.g., TNF-α, IL-8, MCP-1, etc.) are produced in excess, osteoclasts are activated, and activated osteoclasts promote bone resorption. Therefore, suppressing the expression of inflammatory cytokines is important.

[0022] Furthermore, PAI-I is a type of cytokine produced by mast cells and is known to be involved in the progression of thrombosis. It is also known that diabetic patients are more prone to osteoporosis than healthy individuals. In diabetic conditions, increased PAI-I expression in the liver leads to increased blood PAI-I levels, which in turn causes osteoporosis by suppressing osteoblast differentiation or promoting adipogenesis in bone tissue. Therefore, suppressing PAI-1 expression is considered important.

[0023] Furthermore, although not a cytokine, the compositions of this 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). PGs (especially PGE2) activate osteoclasts and promote bone resorption. Therefore, suppressing COX-2 expression is important.

[0024] It is known that moderate activation of PPARγ reduces the size of adipocytes, decreases the expression or secretion of insulin-resistance-inducing cytokines such as TNF-α, and increases the expression or secretion of insulin-sensitizing cytokines such as adiponectin. Therefore, it can be said that upregulation of PPARγ expression is important.

[0025] Adiponectin is known to have a wide range of health benefits, including enhanced insulin sensitivity, suppression of arteriosclerosis, and anti-inflammatory effects. Therefore, it can be said that increasing adiponectin expression is important.

[0026] Examples of animals that may ingest the compositions of this disclosure include humans and other mammals (e.g., rats, mice, rabbits, cattle, pigs, dogs, cats, sheep, monkeys, etc.). Examples of individuals who may require this treatment include: people with increased bone resorption, such as those whose bone metabolism is shifted towards bone resorption due to aging or menopause; patients with osteoporosis or suspected osteoporosis; people with low bone mass; patients with diabetes (high blood sugar) or prediabetes (high blood sugar levels); people with poor dietary habits that are thought to impair the balance of inflammatory cytokines in the body; people with smoking or excessive drinking habits; people who do not exercise regularly and have low levels of physical activity; people with calcium or vitamin D deficiency; and people who feel fatigued on a daily basis and are thought to impair the balance of inflammatory cytokines in the body. Furthermore, even in cases where no major abnormalities are observed in bone metabolism, it can be used preventively to maintain normal bone metabolism.

[0027] The amount of lactic acid bacteria ingested is not particularly limited, but for example, 1 to 100 billion per day (especially per day for adults) is preferred, and 1 to 10 billion per day is more preferred. The amount of lactic acid bacteria contained in the composition disclosed herein can also be set based on the daily intake of lactic acid bacteria. Typically, 1.0 × 10 per gram of dead lactic acid bacteria powder. 11 ~1.0~10 13Since it contains approximately [number] lactic acid bacteria, for example, the composition can be prepared by setting the amount of dead lactic acid bacteria powder in the composition to match the above-mentioned daily intake of lactic acid bacteria, using this value as a reference. 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 of lactic acid bacteria, and the composition can be consumed several times a day (2 or 3 times).

[0028] In this specification, the term "comprising" includes both "consisting essentially of" and "consisting of." Furthermore, this disclosure encompasses all combinations of the constituent elements described herein.

[0029] Furthermore, the various characteristics (properties, structure, function, etc.) described in each embodiment of this disclosure above may be combined in any way to identify the subject matter covered by this disclosure. In other words, this disclosure covers all subject matter consisting of any combination of the combinable characteristics described herein. [Examples]

[0030] The embodiments of this disclosure will be described in more detail below with examples, but the embodiments of this disclosure are not limited to the examples below.

[0031] Animals used Three-week-old male mice (C57BL / 6J (SPF)) were purchased from Charles River Japan Co., Ltd. and used in the experiment. The acclimatization period was from arrival to the day of group division. The quarantine period was set to 7 days, with the arrival day being day 0. During the quarantine and acclimatization period and the test period for group A, D12450J (normal feed (feed 1), RESEARCH DIETS, INC., lot number: 20080303) was placed on the cage lid, and during the test periods for groups B and C, D12492 (high-fat diet (HFD; feed 2), RESEARCH DIETS, INC., lot numbers: 20090202, 20110303) was placed, and the mice were allowed to feed freely except during fasting periods. Drinking water was freely provided throughout the test period using a polysulfone water dispenser (stainless steel tip).

[0032] Grouping Based on body weight measured on the last day of the quarantine and acclimatization period, the animals were divided into groups using a stratified continuous randomization method according to the table below.

[0033] [Table 1]

[0034] The test substance was Lactiplantibacillus. Genus lactic acid bacteria The S25 strain (receipt number: NITE ABP-03571) (dead bacteria) was used. In the table above, the dose is shown in mg / mice, but in terms of bacterial count, it corresponds to 1 billion dead bacteria / day / mice ingested. Distilled water containing the test substance was administered via a tube. Mice in the group not administered the test substance were administered distilled water via a tube. 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 sides of the maxilla, right side of the mandible) and adipose tissue. After cutting the tissues in a frozen state with a scalpel, total RNA was collected using the Total RNA Mini Kit (Bio-Rad), and cDNA was created by reverse transcription. The changes in the expression of the following genes were then observed by real-time PCR. The gene expression results in gingival tissue are shown in Figures 1A-E, and the gene expression results in adipose tissue are shown in Figures 2A-C, as relative values ​​with the gene expression level in the group fed normal diet (Group A: control) set to 1. • TNF-α (tumor necrosis factor-α): gingival tissue • COX-2 (cyclooxygenase-2): Gingival tissue • IL-8 (Interleukin-8): Gingival tissue PAI-1 (plasminogen activator inhibitor-1) Hibita-1): Gingival tissue • MCP-1 (monocyte chemoattractant protein-1): gingival tissue and adipose tissue ·PPARγ (Peroxisome Proliferator-Activated Receptor γ): Adipose tissue • Adiponectin: Adipose tissue

[0035] As shown in Figures 1A-E, Lactiplantibacillus Genus lactic acid bacteria In the group administered with the S25 strain, a reduction in the gene expression levels of TNF-α, COX-2, IL-8, PAI-1, and MCP-1 was observed in gingival tissue. Furthermore, as shown in Figure 2A, it was confirmed that the gene expression level of MCP-1 increased with the intake of a high-fat diet. In adipose tissue, inflammation progresses as obesity progresses and adipocytes enlarge. It has been reported that in obese adipose tissue, increased production of MCP-1 induces macrophage infiltration into the adipose tissue, and the interaction between macrophages and adipocytes worsens the inflammatory state of the adipose tissue, leading to an increase in (macrophage-derived) TNF-α. Since an increase in the gene expression level of MCP-1 was confirmed in adipose tissue, it is thought to be an early inflammatory state. In addition, in gingival tissue, an increase in the gene expression level of TNF-α in addition to MCP-1 was confirmed with the intake of a high-fat diet, suggesting that the inflammatory state may be more advanced than in adipose tissue (systemically). Furthermore, as shown in Figures 2B and 2C, a decrease in the expression levels of PPARγ and adiponectin was confirmed in adipose tissue with the intake of a high-fat diet, and lactiplantibacillus Genus lactic acid bacteria In the group administered with the S25 strain, the decrease in the expression level was suppressed.

[0036] The number of osteoclasts was evaluated using TRAP staining (Acid Phosphatase, Leukocyte (TRAP) Kit (Sigma 387A)). Specifically, slides were immersed in fixative (4% formaldehyde), left to stand at room temperature for 30 seconds, and then washed with distilled water warmed to 37°C. 0.5 ml of Fast GBC Base Solution and 0.5 ml of Sodium Nitrite Solution were placed in a tube and mixed by inversion for 30 seconds, then allowed to stand for 2 minutes. The staining solution was prepared in a beaker, transferred to a staining bottle, warmed to 37°C in a constant temperature bath, and the slides were immersed in the staining bottle, protected from light, and incubated at 37°C for 1 hour. The slides were washed with distilled water and hematoxylin. The samples were immersed in Solution for 2 minutes for counterstaining. They were rinsed with pure water for several minutes, air-dried, and then mounted using glycerol gelatin (Sigma,GG1). They were observed under a microscope, and photographs were taken of the periodontal tissue surrounding the tooth. The number of osteoclasts present near the alveolar bone around the tooth was counted. The number of osteoclasts is shown in Figure 3A, and the TRAP staining results are shown in Figure 3B.

[0037] As shown in Figures 3A and 3B, an increase in the number of osteoclasts was confirmed with high-fat diet intake, and lactiplantibacillus Genus lactic acid bacteria In the group treated with the S25 strain, this increase was suppressed.

Claims

1. A bone resorption inhibitory composition containing Lactiplantibacillus strain S25 (receipt number: NITE ABP-03571).

2. The composition according to claim 1, for use in suppressing osteoclasts.

3. The composition according to claim 1 or 2, which is an oral composition.

4. The composition according to any one of claims 1 to 3, which is a food or beverage composition or a pharmaceutical composition.

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