Composition for preventing or treating arthritis comprising pediococcus acidilactici SY23 strain

Abstract

The present invention relates to a composition comprising a Pediococcus acidilactici SY23 strain (Accession number KCCM 13473P) or a culture thereof as an active ingredient for preventing, alleviating, or treating arthritis.

Description

Composition for preventing or treating arthritis comprising Pediococcus acidilactici SY23 strain

[0001] The present invention relates to a composition for preventing or treating arthritis comprising Pediococcus acidilactici SY23 strain.

[0002] Arthritis is a disease characterized by destruction of joint cartilage and inflammatory changes in the joints. Inflammatory changes within the joints can cause systemic joint swelling and pain, and if exacerbated, can lead to serious consequences, including joint deformities and difficulty bending.

[0003] The direct cause of arthritis remains unclear, and various treatments, including steroids, non-steroidal anti-inflammatory drugs, gout suppressants, and immunosuppressants, are used to treat arthritis. However, these treatments lack fundamental therapeutic effects and have various side effects, limiting their use. Furthermore, some chemotherapeutic agents have limitations, such as limited efficacy in treating existing arthritis.

[0004] Therefore, there is a need to develop an arthritis treatment that is effective in relieving inflammation and pain, has minimal side effects, and is easy to take.

[0005] The present invention provides a pharmaceutical composition for preventing or treating arthritis, which comprises Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) or a culture solution thereof as an active ingredient.

[0006] However, the technical problems to be solved by the present invention are not limited to the problems mentioned above, and other problems not mentioned will be clearly understood by those skilled in the art from the description below.

[0007] The present invention provides a pharmaceutical composition for preventing or treating arthritis, comprising Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) or a culture solution thereof as an active ingredient.

[0008] The above arthritis may include one or more selected from the group consisting of degenerative arthritis, osteoarthritis, rheumatoid arthritis, ankylosing spondylitis, reactive arthritis, psoriatic arthritis, systemic lupus erythematosus, polymyositis, and polymyalgia rheumatica.

[0009] The above arthritis may occur in one or more sites selected from the group consisting of the knees, fingers, jaw, hips, and lower back.

[0010] The pharmaceutical composition can: i) reduce the expression of one or more genes selected from the group consisting of MMP3, MMP9 and MMP13 in cartilage tissue; and ii) reduce the gene expression of IL-1β in cartilage tissue.

[0011] The pharmaceutical composition can change the community of arthritis-related gut microorganism families selected from the group consisting of Bateroidaceae, Lactobacillaceae, Erysipelotrichaceae and Bifidobacteriaceae.

[0012] The pharmaceutical composition can i) increase the population of intestinal microbial species of Bacteroides vulgatus or Clostridium celatum decreased due to arthritis; and ii) decrease the population of intestinal microbial species of Faecalibaculum rodentium, Bacteriodes acidifaciens, Bacteroides stercorirosoris, and Enterorhabdus caecimuris increased due to arthritis.

[0013] The capacity of the above strain is 1×10 5 CFU / day to 1×10 12 It could be CFU / day.

[0014] As one embodiment of the present invention, a health functional food composition for preventing or improving arthritis is provided, which comprises Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) or a culture solution thereof as an active ingredient.

[0015] The Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) according to the present invention is a strain having stable and safe probiotic properties, and the strain (accession number KCCM 13473P) or a culture medium thereof can effectively suppress pain and inflammation in an arthritis (particularly, degenerative arthritis / osteoarthritis) induced model, reduce arthritis-related gene expression, and change the community of arthritis-related intestinal microbial families and species. Therefore, the strain can be usefully utilized as a composition for preventing, improving, or treating arthritis (particularly, degenerative arthritis / osteoarthritis).

[0016] Figure 1 is a diagram showing the experimental design using an animal model of degenerative arthritis / osteoarthritis induced by MIA.

[0017] Figure 2 is a graph showing the changes in the average values ​​of time and force (i.e., 50% avoidance response) for mechanical stimulation of the hind paw according to P. acidilacticiSY23 treatment.

[0018] Figure 3 is a graph showing changes in hind paw weight bearing according to P. acidilacticiSY23 treatment.

[0019] Figure 4 is a photograph and graph showing knee joint inflammation and diameter changes according to P. acidilacticiSY23 treatment.

[0020] Figure 5 is a photograph and graph (H&E) histologically analyzed knee joint tissue according to P. acidilacticiSY23 treatment.

[0021] Figure 6 is a photograph and graph (Safranin-O) showing histological analysis of knee joint tissue following P. acidilacticiSY23 treatment.

[0022] Figure 7 is a graph showing the mRNA expression of cartilage damage-related genes (MMP3, MMP9, MMP13) and inflammation-related genes (IL-1β) in knee joint tissue according to P. acidilacticiSY23 treatment.

[0023] Figure 8 is a graph showing the change in Simpson index in fecal samples according to P. acidilacticiSY23 treatment.

[0024] Figure 9 is a graph showing the change in the Bray-Curtis index in fecal samples according to P. acidilacticiSY23 treatment.

[0025] Figure 10 is a graph showing changes in the community of arthritis-related intestinal microorganisms at the family level in fecal samples according to P. acidilacticiSY23 treatment.

[0026] Figure 11 is a graph showing changes in the community of arthritis-related intestinal microorganisms at the species level in fecal samples according to P. acidilacticiSY23 treatment.

[0027] The present inventors have isolated and identified Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) having a unique metabolic ability among Pediococcus acidilactici strains having probiotic properties, and have completed the present invention by specifically proving its efficacy in treating degenerative arthritis / osteoarthritis in vivo.

[0028]

[0029] Hereinafter, the present invention will be described in detail.

[0030]

[0031] The present invention provides a pharmaceutical composition for preventing or treating arthritis, comprising Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) or a culture solution thereof as an active ingredient.

[0032] In addition, the present invention provides a health functional food composition for preventing or improving arthritis, which comprises Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) or a culture solution thereof as an active ingredient.

[0033]

[0034] The above strain (accession number KCCM 13473P) can be regarded as a subspecies of Pediococcus acidilactici. The above strain (accession number KCCM 13473P) was isolated from traditional fermented foods such as kimchi, and among Pediococcus acidilactici strains, it has a unique metabolic ability and is particularly effective for joint pain and / or inflammation.

[0035] As a result of analyzing 16S rRNA to identify the above-mentioned isolated strain, it was confirmed to have a DNA base sequence encoding 16S rRNA of sequence number 1, and it was deposited with the Biological Resource Center of the Korea Research Institute of Bioscience and Biotechnology on April 24, 2024 and assigned the accession number KCCM 13473P.

[0036] The above strain (accession number KCCM 13473P) can be produced as a live cell or a dead cell. In addition, the culture medium can be a culture filtrate, i.e., a supernatant from which the cell has been removed.

[0037] The capacity of the above strain (accession number KCCM 13473P) is 1×10 5 CFU / day to 1×10 12 CFU / day can be 1×10 8 CFU / day to 1×10 10 CFU / day is preferred, but is not limited to this.

[0038]

[0039] The term “arthritis” in this specification refers to a disease in which joint cartilage is destroyed, inflammatory changes occur in the joint, and joint swelling and pain are caused as a result.

[0040] The above arthritis is a broad concept including at least one selected from the group consisting of degenerative arthritis, osteoarthritis, rheumatoid arthritis, ankylosing spondylitis, reactive arthritis, psoriatic arthritis, systemic lupus erythematosus, polymyositis, and polymyalgia rheumatica, and is preferably degenerative arthritis or osteoarthritis, but is not limited thereto.

[0041] Additionally, the arthritis may occur in one or more sites selected from the group consisting of the knees, fingers, jaw, hips, and lower back.

[0042]

[0043] First, the strain (accession number KCCM 13473P) or its culture solution can effectively suppress pain and inflammation and reduce arthritis-related gene expression in an arthritis (particularly, degenerative arthritis / osteoarthritis)-induced model. At this time, arthritis (particularly, degenerative arthritis / osteoarthritis)-induced model can be induced by injecting monosodium iodoacetate (MIA) into the joint cavity of an animal model.

[0044] Specifically, the pharmaceutical composition can i) reduce the expression of cartilage damage-related genes (particularly, one or more genes selected from the group consisting of MMP3, MMP9 and MMP13) in cartilage tissue; and ii) reduce the expression of inflammation-related genes (particularly, the gene for IL-1β) in cartilage tissue.

[0045]

[0046] Next, first, the strain (accession number KCCM 13473P) or its culture medium can change the community of arthritis-related intestinal microorganism families and species in an arthritis (particularly, degenerative arthritis / osteoarthritis)-induced model. At this time, the arthritis (particularly, degenerative arthritis / osteoarthritis)-induced model can induce arthritis by injecting monosodium iodoacetate (MIA) into the joint space of an animal model. In addition, by collecting and analyzing a fecal sample of the arthritis (particularly, degenerative arthritis / osteoarthritis)-induced model, the change in the community of arthritis-related intestinal microorganisms can be measured.

[0047] Specifically, the pharmaceutical composition can change the community of arthritis-related gut microorganism families selected from the group consisting of Bateroidaceae, Lactobacillaceae, Erysipelotrichaceae, and Bifidobacteriaceae.

[0048] In addition, the pharmaceutical composition can i) increase the population of intestinal microbial species of Bacteroides vulgatus or Clostridium celatum decreased due to arthritis; and ii) decrease the population of intestinal microbial species of Faecalibaculum rodentium, Bacteriodes acidifaciens, Bacteroides stercorirosoris, and Enterorhabdus caecimuris increased due to arthritis.

[0049]

[0050] The pharmaceutical composition according to the present invention can be formulated and used in the form of oral preparations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., external preparations, suppositories, and sterile injection solutions, according to conventional methods, and may include appropriate carriers, excipients, or diluents conventionally used in the manufacture of pharmaceutical compositions for formulation.

[0051] The carrier or excipient or diluent may include various compounds or mixtures including lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate and mineral oil.

[0052] When formulating, it can be manufactured using diluents or excipients such as fillers, weighting agents, binders, wetting agents, disintegrants, and surfactants that are commonly used.

[0053] Solid preparations for oral administration can be prepared by mixing the strain (accession number KCCM 13473P) with at least one excipient, such as starch, calcium carbonate, sucrose or lactose, or gelatin. In addition to simple excipients, lubricants such as magnesium stearate and talc can also be used.

[0054] Liquid preparations for oral administration include suspensions, solutions, emulsions, and syrups, and in addition to commonly used simple diluents such as water and liquid paraffin, they may contain various excipients such as wetting agents, sweeteners, fragrances, and preservatives.

[0055] Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, and suppositories. Non-aqueous solutions and suspensions include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. Suppository bases include witepsol, macrogol, Tween 61, cocoa butter, laurin, and glycerol gelatin.

[0056] The preferred dosage of the pharmaceutical composition according to the present invention varies depending on the patient's condition, body weight, severity of the disease, drug form, route of administration, and duration of administration, but can be appropriately selected by those skilled in the art. However, for desirable effects, the dosage may be 0.0001 to 2,000 mg / kg per day, preferably 0.001 to 2,000 mg / kg. Administration may be once a day or divided into several doses. However, the scope of the present invention is not limited by the above dosage.

[0057] The pharmaceutical composition according to the present invention can be administered to mammals such as rats, mice, livestock, and humans via various routes. All modes of administration can be administered, for example, orally, rectally, or by intravenous, intramuscular, subcutaneous, intrauterine, or intracerebroventricular injection.

[0058]

[0059] In the health functional food composition according to the present invention, when the strain (accession number KCCM 13473P) is used as an additive to a health functional food, it can be added as is or used together with other foods or food ingredients, and can be used appropriately according to conventional methods. The amount of active ingredients mixed can be appropriately determined depending on the intended use, such as prevention, health, or treatment.

[0060] The formulation of health functional foods can be in the form of powders, granules, pills, tablets, capsules, or any other form of general food or beverage.

[0061] There is no particular limitation on the types of the above foods, and examples of foods to which the above substances can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes, and all foods in the conventional sense can be included.

[0062] In general, when manufacturing food or beverages, the strain (accession number KCCM 13473P) may be added in an amount of 15 parts by weight or less, preferably 10 parts by weight or less, per 100 parts by weight of the raw material. However, in the case of long-term consumption for health and hygiene purposes or health control purposes, the amount may be below the above range.

[0063] Among the health functional foods according to the present invention, the beverage may contain various flavoring agents or natural carbohydrates as additional ingredients, just like conventional beverages. The natural carbohydrates mentioned above may be 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. As a sweetener, a natural sweetener such as thaumatin or stevia extract, or a synthetic sweetener such as saccharin or aspartame may be used. The proportion of the natural carbohydrate may be about 0.01 to 0.04 g, preferably about 0.02 to 0.03 g, per 100 mL of the beverage according to the present invention.

[0064] In addition to the above, the health functional food composition according to the present invention may contain various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, and carbonating agents used in carbonated beverages. In addition, the health functional food composition according to the present invention may contain fruit pulp for the production of natural fruit juice, fruit juice drinks, and vegetable drinks. These ingredients may be used independently or in mixtures. The ratio of these additives is not limited, but is generally selected in the range of 0.01 to 0.1 parts by weight relative to 100 parts by weight of the health functional food composition according to the present invention.

[0065]

[0066] Alternatively, the present invention provides a use for a composition for preventing, treating or improving arthritis, which comprises Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) or a culture solution thereof as an active ingredient.

[0067] Alternatively, the present invention provides a method for preventing, treating, or improving arthritis, comprising Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) or a culture solution thereof as an active ingredient. In this case, the term "subject" refers to a subject requiring treatment for a disease, and more specifically, refers to mammals such as humans or non-human primates, mice, rats, dogs, cats, horses, and cows.

[0068]

[0069] As described above, the Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) according to the present invention is a strain having stable and safe probiotic properties, and the strain (accession number KCCM 13473P) or a culture medium thereof can effectively suppress pain and inflammation in an arthritis (particularly, degenerative arthritis / osteoarthritis) induced model, reduce arthritis-related gene expression, and change the community of arthritis-related intestinal microbial families and species. Therefore, the strain can be usefully utilized as a composition for preventing, improving, or treating arthritis (particularly, degenerative arthritis / osteoarthritis).

[0070]

[0071] Hereinafter, preferred examples are presented to aid in understanding the present invention. However, the following examples are provided solely to facilitate a better understanding of the present invention, and the scope of the present invention is not limited by the following examples.

[0072]

[0073] <Example>

[0074] Example 1: Isolation and identification of P. acidilacticiSY23 strain from kimchi

[0075] <1-1> Pure isolation of kimchi-derived microorganisms

[0076] In order to carry out an embodiment of the present invention, microorganisms were purified from kimchi. Specifically, kimchi broth such as cabbage kimchi, radish kimchi, and cubed radish kimchi was separated into 10 parts by distilling water. -1 10 inland -7Kimchi samples were prepared by serially diluting them in a ratio of . Then, the kimchi samples were spread on Pediococcus selective medium (PES) agar, a lactic acid bacteria selective medium, and isolated. The spread medium was cultured at 37℃, and the resulting colonies were collected and observed for morphology. Then, the colonies were streaked on new medium several times and the microorganisms were purified.

[0077] <1-2> Identification of isolated lactic acid bacteria

[0078] In order to confirm the strain of the microorganism isolated through the lactic acid bacteria selection medium, the isolated microorganism was identified.

[0079] Specifically, genomic DNA was extracted from the isolated strain and sequenced (by Bionics, Korea) to obtain the 16S rRNA gene sequence. The 16S rRNA sequence was analyzed for homology with other standard strains registered in the database using the BLAST program, and strains showing more than 98% homology were identified. In addition, homology was analyzed and a phylogenetic tree was constructed using the Clustal X and Mega 7 programs.

[0080] As a result, the lactic acid bacteria isolated from kimchi showed homology with strains reported as Pediococcus acidilactici, and was named Pediococcus acidilacticiSY23 strain, and deposited at the Korea Center for Microorganism Conservation (KCCM) on April 24, 2024 (accession number KCCM 13473P).

[0081]

[0082] Example 2: Experimental design using an animal model of degenerative arthritis / osteoarthritis induced by MIA.

[0083] 57BL / 6N male, 8-week-old mice were prepared. After removing hair around the left knee of the anesthetized mice, 10 μL (100 mg / mL) of monosodium iodoacetate (MIA) was injected into the knee joint space to induce degenerative arthritis / osteoarthritis. L. brevisKU15122 live cells, P. acidilacticiSY21 (accession number KCCM 11614P) live cells, and P. acidilacticiSY23 live cells were each injected at a density of 2 × 10 per mouse per day. 9 CFU concentrations were administered orally daily for a total of 6 weeks, from 2 weeks before to 4 weeks after induction of degenerative arthritis / osteoarthritis (Table 1 and Figure 1).

[0084] Group Treatment Method Notes None Vehicle Control PBS (MIA + PBS or MIA) MIA + PBS Arthritis-induced group Indo (MIA + Indo) MIA + indomethacine (2 mg / kg) MIA and treatment-treated control group KU15122 MIA + L. brevis KU15122 MIA and L. brevis KU15122-treated comparison group SY21 MIA + P. acidilactici SY21 MIA and P. acidilactici SY21-treated comparison group SY23 (MIA + SY23) MIA + P. acidilactici SY23 MIA and P. acidilactici SY23-treated experimental group

[0085]

[0086] Example 3: Measurement of pain changes following P. acidilacticiSY23 treatment in an animal model of degenerative arthritis / osteoarthritis.

[0087] First, a paw withdrawal threshold (PWT) experiment was conducted to assess pain thresholds for mechanical stimulation. Mechanical stimulation was applied to the hind paws of mice with and without osteoarthritis / degenerative arthritis, and the paw withdrawal response was assessed. The time and force required to withstand the mechanical stimulation for each hind paw were measured five times, and the average value was designated as a 50% withdrawal response (Fig. 2).

[0088] As shown in Fig. 2, the 50% avoidance response was significantly reduced in the group in which only arthritis was induced (PBS), but the MIA and P. acidilacticiSY23-treated experimental group (SY23) significantly increased the values ​​of the time and force required to withstand mechanical stimulation, thereby increasing the 50% avoidance response, which was confirmed to be at a similar level to the MIA and treatment-treated control group (Indo).

[0089] Additionally, a hind paw weight-bearing test was performed. Three weeks after intra-articular injection of MIA into the knee joint of mice, the body weights of hind paws with and without degenerative arthritis / osteoarthritis were measured five times each. The average value obtained by dividing the values ​​by body weight was defined as the hind paw weight-bearing capacity (Fig. 3).

[0090] As shown in Fig. 3, it was confirmed that the weight bearing of the hind paw was significantly recovered in the MIA and P. acidilacticiSY23 treatment experimental group (SY23) compared to the group in which only arthritis was induced (PBS).

[0091]

[0092] Example 4: Measurement of inflammatory changes following P. acidilacticiSY23 treatment in an animal model of degenerative arthritis / osteoarthritis.

[0093] First, to observe the degree of inflammation in the knee joint area 3 weeks after MIA injection into the knee joint cavity of mice, the MMPsense-680 probe (PerkinElmer, 2 nmol / 150 μL in PBS) was injected intravenously into the blood vessel, and the fluorescent signal was observed at the area of ​​arthritis induced through in vivo imaging. In addition, to confirm whether inflammation was induced in the knee joint cavity of mice, the change in diameter between the outer and inner sides of the knee joint was measured using calipers before and after MIA injection, and the degree of damage to the cartilage and synovial area was observed (Fig. 4).

[0094] As shown in Fig. 4, the change in knee joint diameter in the MIA and P. acidilacticiSY23-treated experimental group (SY23) was significantly reduced compared to the arthritis-induced group (PBS), indicating a significant reduction in the extent of damage to the cartilage and synovial membrane. This can be considered effective compared to the MIA and treatment-treated control group (Indo).

[0095]

[0096] Example 5: Measurement of histopathological changes following P. acidilacticiSY23 treatment in an animal model of degenerative arthritis / osteoarthritis.

[0097] After sacrificing the mice, the knee area was cut and stored in a 10% chloroform solution. After decalcification of the knee joint tissue in a solution containing 10% EDTA, the knee joint tissue was embedded in paraffin wax and fixed. The knee joint tissue was sectioned to a thickness of 7 μm, stained with H&E and Safranin-O, and the condition of the tissue was observed using a light microscope at a magnification of ×100 (Figs. 5 and 6).

[0098] As shown in Fig. 5, when the knee joint tissue was confirmed through analysis of the H&E staining results, in the group in which only arthritis was induced (PBS), degenerative arthritis / osteoarthritis was induced, and compared to the vehicle control group (Con), fibrous tissue deformation and inflammatory cell infiltration were significantly observed in the cartilage and synovial areas. However, when treated with live cells, especially in the MIA and P. acidilacticiSY23 treated experimental group (SY23), fibrous tissue deformation and inflammatory cell infiltration in the cartilage and synovial areas were significantly reduced. This can be considered effective compared to the MIA and treatment control group (Indo).

[0099] As shown in Fig. 6, the damage to cartilage tissue was confirmed through analysis of the Safranin-O staining results that stain the proteoglycan layer. As a result, in the group that only induced arthritis (PBS), degenerative arthritis / osteoarthritis was induced, and significant deformation of joint tissue and damage to cartilage cells around the synovium were confirmed compared to the vehicle control group (Con). However, when treated with live bacteria, especially in the MIA and P. acidilacticiSY23 treated experimental group (SY23), deformation of joint tissue was reduced, and damage to cartilage cells was also confirmed to be reduced. This can be considered effective compared to the MIA and treatment control group (Indo).

[0100]

[0101] Example 6: Measurement of Arthritis-Related Gene Expression Following P. acidilacticiSY23 Treatment in an Animal Model of Degenerative Arthritis / Osteoarthritis

[0102] After sacrificing the mice, knee joint tissues were isolated and the mRNA expression of genes related to cartilage damage and inflammation was compared.

[0103] Knee joint tissue was transferred to a pre-filled bead tube (Lysing Matrix E, MP Biomedicals, CA, USA), and 1 mL of TRIzol reagent (Invitrogen, MA, USA) was added per 50-100 mg of tissue. The tissue was beat twice for 40 seconds using a bead beater (BEADBLASTER 24, Benchmark Scientific, NJ, USA). The tissue was left to stand at room temperature for 5 minutes, and 0.2 mL of chloroform per 1 mL of TRIzol was added and centrifuged at 4°C, 12,000 rpm, and for 15 minutes. 0.5 mL of isopropanol was added to the supernatant, and centrifuged at 4°C, 12,000 rpm, and for 10 minutes. The supernatant was removed, and 1 mL of 75% (v / v) ethanol was added and centrifuged for 5 minutes. After removing the supernatant and drying for 30 minutes to 1 hour, RNA was dissolved in 0.2-0.3 mL of nuclease-free water (Noble Bio, Gyeonggi, Korea) and used.

[0104] RNA concentration was measured using Nanodrop 2000 (Thermo Fisher Scientific, USA), and cDNA was synthesized using a cDNA synthesis kit (TOPscriptTM, Daejeon, Korea) using 1 ng of RNA. The synthesized cDNA was subjected to real-time quantitative PCR (Bio-Rad CFX-96, CA, USA) analysis using TOPreal qPCR 2× premix (Enzynomics, Korea). The primer sequences are shown in Table 2 below (Fig. 7).

[0105] Gene sequence MMP3 Forward CTC TGG AAC CTG AGA CAT CAC C Reverse AGG AGT CCT GAG AGA TTT GCG CMMP9 Forward GCT GAC TAC GAT AAG GAC GGC A Reverse TAG TGG TGC AGG CAG AGT AGG AMMP13 Forward TTC TGG TCT TCT GGC ACA CGC TTT Reverse CCA AGC TCA TGG GCA GCA ACA ATAIL-1β Forward ATG CCA CCT TTT GAC AGT GAT GG Reverse AGC TTC TCC ACA GCC ACA ATG APDH Forward GTG TTC CTA CCC CCA ATG TGT Reverse ATT GTC ATA CCA GGA AAT GAG CTT

[0106] As shown in Fig. 7, in the group in which only arthritis was induced (PBS), compared to the vehicle control group (Con), the mRNA expression of cartilage damage-related genes (MMP3, MMP9, MMP13) and inflammation-related genes (IL-1β) in the cartilage and synovial membrane areas increased, but when treated with live cells, especially in the MIA and P. acidilacticiSY23-treated experimental group (SY23), the mRNA expression of cartilage damage-related genes (MMP3, MMP9, MMP13) and inflammation-related genes (IL-1β) was significantly reduced.

[0107]

[0108] Example 7: Measurement of changes in the gut microbiota community associated with arthritis following P. acidilacticiSY23 treatment in an animal model of degenerative arthritis / osteoarthritis.

[0109] To measure changes in the gut microbiota community associated with arthritis, fecal samples were collected from mice, and sequence reads were obtained after quality filtering (Figs. 8 to 10).

[0110] As shown in Fig. 8, the Simpson index, which indicates the uniformity among the species of microorganisms existing in the same group, showed a significant difference in all groups (MIA+PBS, MIA+Indo, MIA+SY23) in which degenerative arthritis / osteoarthritis was induced compared to the vehicle control group (Con).

[0111] As shown in Fig. 9, the Bray-Curtis index, which indicates changes in the intestinal microbiota community, was confirmed to have different clusters in all groups (MIA+PBS, MIA+Indo, MIA+SY23) in which degenerative arthritis / osteoarthritis was induced compared to the vehicle control group (Con).

[0112] At the Phylum level, the differences in the clusters between all groups (MIA+PBS, MIA+Indo, MIA+SY23) in which degenerative arthritis / osteoarthritis was induced compared to the control group (Con) were all minimal. However, as shown in Fig. 10, at the Family level, differences in the clusters of intestinal microorganisms of the Bateroidaceae, Lactobacillaceae, Erysipelotrichaceae, and Bifidobacteriaceae families were confirmed in all groups (MIA+PBS, MIA+Indo, MIA+SY23) in which degenerative arthritis / osteoarthritis was induced compared to the control group (Con). Meanwhile, at the genus level, differences in the communities of various intestinal microorganisms were confirmed in all groups (MIA+PBS, MIA+Indo, MIA+SY23) in which degenerative arthritis / osteoarthritis was induced compared to the control group (Con).

[0113] Meanwhile, at the species level, linear discriminant analysis effect size (LEfSe) analysis was used to identify gut microbial species that exhibited an LDA score of 2 or higher, which characterizes cluster differences (Fig. 11).

[0114] As shown in Fig. 11, the MIA and P. acidilacticiSY23 treatment experimental group (SY23) was confirmed to have increased the intestinal microbial species community of Bacteroides vulgatus and Clostridium celatum, which were reduced in the arthritis-only induced group (MIA). Meanwhile, MIA and P. In the acidilacticiSY23 treatment group (SY23), it was confirmed that the population of intestinal microbial species, Faecalibaculum rodentium, Bacteriodes acidifaciens, Bacteroides stercorirosoris, and Enterorhabdus caecimuris, which were increased in the arthritis-only induced group (MIA), was significantly reduced again.

[0115]

[0116] Below, an example of a composition containing the strain or culture medium of the present invention as an effective ingredient is described, but the present invention is not intended to be limited thereto, but is merely intended to be described in detail.

[0117]

[0118] Preparation Example 1: Preparation of a powder

[0119] 20 mg of Pediococcus acidilacticiSY23 strain or culture

[0120] 100 mg of lactose monohydrate

[0121] 10 mg of talc

[0122] The above ingredients were mixed and filled into a sealed bag to prepare a powder.

[0123]

[0124] Preparation Example 2: Preparation of tablets

[0125] 10 mg of Pediococcus acidilacticiSY23 strain or culture

[0126] 100 mg of corn starch

[0127] 100 mg of lactose monohydrate

[0128] 2 mg of magnesium stearate

[0129] After mixing the above ingredients, tablets were manufactured by pressing them according to a conventional tablet manufacturing method.

[0130]

[0131] Preparation Example 3: Preparation of capsules

[0132] 10 mg of Pediococcus acidilacticiSY23 strain or culture

[0133] 3 mg of microcrystalline cellulose

[0134] 14.8 mg of lactose monohydrate

[0135] Magnesium stearate 0.2 mg

[0136] After mixing the above ingredients, the mixture was filled into a gelatin capsule according to a conventional capsule manufacturing method to produce a capsule.

[0137]

[0138] Preparation Example 4: Preparation of Injection

[0139] 10 mg of Pediococcus acidilacticiSY23 strain or culture

[0140] Mannitol 180 mg

[0141] 2974 mg of sterile distilled water for injection

[0142] 26 mg of sodium dihydrogen phosphate

[0143] After mixing the above ingredients, the composition was prepared with the above ingredient content per 1 ampoule (2 mL) according to the manufacturing method of a conventional injection.

[0144]

[0145] Preparation Example 5: Preparation of a liquid formulation

[0146] 10 mg of Pediococcus acidilacticiSY23 strain or culture

[0147] 10 g of isoflavonoids

[0148] 5 g of mannitol

[0149] Appropriate amount of purified water

[0150] Lemon scent as needed

[0151] The above ingredients are dissolved in purified water according to the usual manufacturing method, an appropriate amount of lemon flavor is added, and then purified water is added to adjust the total volume to 100 mL, sterilize, and fill into a brown bottle to prepare a liquid.

[0152]

[0153] Preparation Example 6: Manufacturing of health functional food

[0154] 10 mg of Pediococcus acidilacticiSY23 strain or culture

[0155] Vitamin mixture appropriate amount

[0156] Vitamin A acetate 70 μg

[0157] Vitamin E 1.0 mg

[0158] Vitamin B1 0.13 mg

[0159] Vitamin B2 0.15 mg

[0160] Vitamin B6 0.5 mg

[0161] Vitamin B12 0.2 μg

[0162] Vitamin C 10 mg

[0163] 10 μg of biotin

[0164] Nicotinamide 1.7 mg

[0165] 50 μg of folic acid

[0166] Calcium pantothenate 0.5 mg

[0167] Appropriate amount of mineral mixture

[0168] 1.75 mg of ferrous sulfate

[0169] 0.82 mg of zinc oxide

[0170] Magnesium carbonate 25.3 mg

[0171] 15 mg of monobasic potassium phosphate

[0172] 55 mg of dibasic calcium phosphate

[0173] Potassium citrate 90 mg

[0174] 100 mg of calcium carbonate

[0175] Magnesium chloride 24.8 mg

[0176] The composition ratio of the above vitamin and mineral mixture is a preferred example of a mixture of ingredients relatively suitable for health functional foods, but the mixing ratio may be arbitrarily modified, and the above ingredients may be mixed according to a conventional health functional food manufacturing method, granules may be manufactured, and used in the manufacture of health functional foods according to a conventional method.

[0177]

[0178] Preparation Example 7: Manufacturing a health drink

[0179] 10 mg of Pediococcus acidilacticiSY23 strain or culture

[0180] 15 g of vitamin C

[0181] Vitamin E (powder) 100 g

[0182] 19.75 g of ferrous lactate

[0183] 3.5 g of zinc oxide

[0184] 3.5 g of nicotinamide

[0185] Vitamin A 0.2 g

[0186] Vitamin B1 0.25 g

[0187] Vitamin B2 0.3g

[0188] Quantity of purified water

[0189] The above ingredients are mixed according to a conventional health drink manufacturing method, then stirred and heated at 85°C for about 1 hour, the resulting solution is filtered, placed in a sterilized 2 L container, sealed and sterilized, and then stored in a refrigerator, and then used to manufacture the health drink composition of the present invention.

[0190]

[0191] The above composition ratio is a preferred example of a mixture of ingredients suitable for a relatively preferred beverage, but the mixing ratio may be arbitrarily modified according to regional or ethnic preferences such as the demand class, demand country, and intended use.

[0192]

[0193] The foregoing description of the present invention is provided for illustrative purposes only. Those skilled in the art will readily appreciate that the present invention can be readily modified into other specific forms without altering the technical spirit or essential characteristics of the present invention. Therefore, the embodiments described above should be understood as illustrative in all respects and not restrictive.

[0194]

[0195] [Accession number]

[0196] Name of depositor: Korea Microbiological Conservation Center (overseas)

[0197] Accession number: KCCM 13473P

[0198] Concession Date: April 24, 2024

[0199]

[0200]

Claims

1. A pharmaceutical composition for preventing or treating arthritis, comprising Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) or a culture solution thereof as an active ingredient.

2. In paragraph 1, A pharmaceutical composition characterized in that the arthritis comprises at least one selected from the group consisting of degenerative arthritis, osteoarthritis, rheumatoid arthritis, ankylosing spondylitis, reactive arthritis, psoriatic arthritis, systemic lupus erythematosus, polymyositis, and polymyalgia rheumatica.

3. In paragraph 1, A pharmaceutical composition, characterized in that the arthritis occurs in one or more areas selected from the group consisting of knees, fingers, jaw, hips, and waist.

4. In paragraph 1, The above pharmaceutical composition i) reducing the expression of one or more genes selected from the group consisting of MMP3, MMP9 and MMP13 in cartilage tissue; ⅱ) A pharmaceutical composition characterized by reducing gene expression of IL-1β in cartilage tissue.

5. In paragraph 1, A pharmaceutical composition characterized in that the pharmaceutical composition changes the community of arthritis-related gut microorganism families selected from the group consisting of Bateroidaceae, Lactobacillaceae, Erysipelotrichaceae, and Bifidobacteriaceae.

6. In paragraph 1, The above pharmaceutical composition i) Increase the population of intestinal microbial species of Bacteroides vulgatus or Clostridium celatum that are reduced due to arthritis; ⅱ) A pharmaceutical composition characterized in that it reduces the population of intestinal microbial species of Faecalibaculum rodentium, Bacteriodes acidifaciens, Bacteroides stercorirosoris, and Enterorhabdus caecimuris increased due to arthritis.

7. In paragraph 1, The capacity of the above strain is 1×10 5 CFU / day to 1×10 12 A pharmaceutical composition characterized by having CFU / day.

8. A health functional food composition for preventing or improving arthritis, comprising Pediococcus acidilactici SY23 strain (accession number KCCM 13473P) or a culture solution thereof as an active ingredient.

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