Streptococcus salivarius CCFM1512, which metabolizes a product that inhibits excessive inflammatory response to relieve periodontitis
By using the fermentation supernatant of Streptococcus salivarius CCFM1512 and metabiotics, the host immune response was regulated, which solved the problem of host immune-dependent inflammation in periodontitis and achieved the relief of periodontitis symptoms and the restoration of tissue structure.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- JIANGNAN UNIV
- Filing Date
- 2026-04-28
- Publication Date
- 2026-07-28
AI Technical Summary
Current treatments for periodontitis mainly focus on eliminating pathogens, but long-term inflammatory responses depend on the host's immune system, leading to systemic inflammation, and there is a lack of effective measures to regulate the host's immunity.
The fermentation supernatant of Streptococcus salivarius CCFM1512 and/or its metagenes were used to alleviate periodontitis by modulating the host immune response and inhibiting excessive inflammatory response.
It reduces the expression of pro-inflammatory factors induced by Porphyromonas gingivalis lipopolysaccharide, alleviates alveolar bone resorption, regulates macrophage polarization balance, restores periodontal tissue structure, inhibits inflammatory factor levels, and promotes inflammation resolution.
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Figure CN122465766A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to a strain of Streptococcus salivans CCFM1512 whose metabolites can inhibit excessive inflammatory responses and alleviate periodontitis, belonging to the field of microbial technology. Background Technology
[0002] Periodontitis is a very common oral disease, mainly characterized by chronic inflammation of the periodontal soft tissues, loss of periodontal attachment, alveolar bone resorption, and ultimately tooth loss. Early epidemiology suggested that *Porphyromonas gingivalis* (…) was the primary cause of tooth loss. Porphyromonas gingivalis ), Actinobacillus consanguineus ( Actinobacillus actinomycetemcornitans ) and Forsythia ( Bacteroides forsythus Gram-negative anaerobic or microaerophilic bacteria, such as [unspecified bacteria], are the main factors leading to and promoting the development of periodontitis. Therefore, the main treatment focuses on eliminating or reducing the bioburden of these pathogens, including subgingival scaling (SRP) and local or systemic antibiotic therapy.
[0003] However, the development of periodontitis largely depends on the host's own immune characteristics. The tissue damage caused by inflammation is entirely due to the host's own immune system, and the peptides and heme iron produced by the breakdown of host tissues continue to provide nutrients for the growth of pathogens. Prolonged and excessive inflammatory responses provide periodontal pathogens and their virulence factors with opportunities to enter the external circulation. This hematogenous translocation leads to systemic inflammation, subsequently inducing a series of diseases such as cardiovascular disease, type II diabetes, osteoporosis, and inflammatory bowel disease. Therefore, timely reversal of the excessive immune response during periodontitis by regulating the host's immune system to promote inflammatory resolution may be an important direction for future relief of periodontitis symptoms. Its effector pathways include regulating the macrophage M1 / M2 polarization balance and regulating DEL-1 expression to inhibit excessive neutrophil recruitment.
[0004] Numerous studies have revealed that certain components of the fermentation supernatant or cells of beneficial bacterial strains have positive regulatory effects on the host's immune response. For example, the surface protein of *Lactobacillus acidophilus* NCFM can alleviate colitis by inducing regulatory T cell differentiation; extracellular vesicles carrying capsular polysaccharides in *Bacteroides fragilis* NCTC9343 can prevent colitis by inducing regulatory T cell differentiation and the production of anti-inflammatory cytokines; and indole-3-lactic acid in the fermentation supernatant of *Lactobacillus plantarum* L168 can alleviate acute colitis by regulating inflammatory cytokine levels. In summary, extending from intestinal mucosal immunity to oral mucosal immunity, identifying oral commensal bacteria and their metabolites that can inhibit excessive inflammatory responses is of great significance for adjuvant treatment of periodontitis. Summary of the Invention
[0005] Therefore, the purpose of this invention is to provide a fermentation supernatant of a strain of Streptococcus salivarius that can alleviate periodontitis by inhibiting excessive inflammatory response.
[0006] This invention provides *Streptococcus salivarius* (Salivarius) Streptococcus salivarius CCFM1512, taxonomically named Streptococcus salivarius It was deposited on August 21, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No: 66863, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou.
[0007] The salicylic streptococcus ( Streptococcus salivarius CCFM1512 is a Gram-positive bacterium. Under a microscope, the cells are oval and exist in single, paired, or chain form. After inoculation on MRS solid medium and cultured for 48 h, the colonies are generally white protrusions with a smooth, moist surface and regular edges.
[0008] The present invention also provides a metagene of *Streptococcus salivarius* CCFM1512, which is prepared by any of the following methods: (a) After fermenting the *Streptococcus salivarius* CCFM1512, collect the fermentation supernatant; (b) After fermenting the said Streptococcus salivarius CCFM1512, collect the bacterial cells and lyse them to obtain the bacteria.
[0009] The present invention also provides a microbial preparation containing the *Streptococcus salivarius* CCFM1512 and / or its postbiotic.
[0010] In one embodiment, the content of *Streptococcus salivarius* CCFM1512 in the microbial preparation is not less than 1 × 10⁻⁶. 6 CFU / mL or 1×10 6 CFU / g.
[0011] The present invention also provides products containing the aforementioned Streptococcus salivarius CCFM1512 and / or its postbiotics, wherein the products are food, health products, pharmaceuticals or daily chemical products. In one embodiment, the food contains the *Streptococcus salivarius* CCFM1512 and / or its postbiotics, as well as conventional excipients.
[0012] In one embodiment, the conventional excipients include fillers, flavoring agents, binders, disintegrants, lubricants, etc. One or more of the following: antacids and nutritional fortifiers.
[0013] In one embodiment, the health product contains the *Streptococcus salivarius* CCFM1512 and / or its postbiotics, as well as conventional excipients.
[0014] In one embodiment, the conventional excipients include fillers, flavoring agents, binders, disintegrants, lubricants, etc. One or more of the following: antacids and nutritional fortifiers.
[0015] In one embodiment, the drug contains the salicylic streptococcus CCFM1512 and / or its postgenes, as well as a pharmaceutically permissible carrier.
[0016] In one embodiment, the pharmaceutically permissible carrier includes one or more of the following commonly used medical fillers, binders, humectants, disintegrants, lubricants, and flavoring agents.
[0017] In one embodiment, the personal care product includes toothpaste, mouthwash, or oral spray.
[0018] In one embodiment, the content of *Streptococcus salivarius* CCFM1512 in the product is not less than 1 × 10⁻⁶. 6 CFU / mL or 1×10 6 CFU / g.
[0019] The present invention also provides the use of Streptococcus salivarius CCFM1512 and / or its metabolites in medicaments for inhibiting oral inflammatory responses and / or alleviating periodontitis.
[0020] Beneficial effects: This invention provides a strain of Streptococcus salivarius (Salivarius) Streptococcus salivarius CCFM1512 has the effect of inhibiting excessive inflammatory response and relieving periodontitis, specifically manifested in: (1) Reduce the expression level of pro-inflammatory factors in human gingival fibroblasts (HGF) induced by Porphyromonas gingivalis lipopolysaccharide (Pg-LPS); (2) Alleviating alveolar bone resorption in a rat model of periodontitis; (3) Reduced the level of inflammatory factors in rat gingival tissue; (4) It alleviates the degradation of collagen fibers in the periodontal tissues of rats; (5) Regulate the expression level of endothelial development site-1 (DEL-1) in rat periodontal tissue; (6) Restore the M1 / M2 polarization balance of macrophages in rat periodontal tissue; Therefore, the fermentation supernatant of Streptococcus salivarius CCFM1512 has great application potential in products that regulate host oral immunity, inhibit excessive inflammatory response, and alleviate periodontitis.
[0021] Preservation of biological materials Salinomyces ( Streptococcus salivarius CCFM1512, taxonomically named Streptococcus salivarius It was deposited on August 21, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No: 66863, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou. Attached Figure Description
[0022] Figure 1 The effect of intervention with Streptococcus salivarius fermentation supernatant on the expression of related inflammatory factors in a Pg-LPS-induced HGF inflammation model.
[0023] Figure 2 Design a flowchart for animal experiments; A is the animal experiment flowchart; B is the process of ligating the second maxillary molar of a rat with silk thread. Figure 3 The effect of intervention with the fermentation supernatant of Streptococcus saliva on alveolar bone resorption in rats; A is the alveolar bone of rats scanned by Micro-CT; B is the distance from the cementoenamel junction (CEJ) to the alveolar ridge crest (ABC) in each group; C is the alveolar bone density in each group.
[0024] Figure 4 The images show the histopathological findings and DEL-1 expression in rat periodontal tissue; A shows HE staining of rat periodontal tissue sections; B shows Masson staining of rat periodontal tissue sections; C shows immunohistochemical staining of DEL-1 protein in rat periodontal tissue sections; D shows DEL-1 expression in rat gingival tissue. EDIL3 The relative expression level of the gene; E represents the IL-17A content in rat gingival tissue; F represents the MPO content in rat gingival tissue.
[0025] Figure 5 The effects of *Streptococcus saliva* fermentation supernatant intervention on macrophage polarization in rat periodontal tissues; A: Immunofluorescence staining of rat periodontal tissue sections; B: IL-1β content in rat gingival tissue; C: IL-6 content in rat gingival tissue; D: IL-6 content in rat gingival tissue. iNOS Relative gene expression levels; E represents the expression level in rat gingival tissue. Arg-1 The relative expression level of genes. Detailed Implementation
[0026] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments and accompanying drawings. The strains, cells, and animals involved in the following embodiments are as follows: SPF-grade male Wistar rats, 6 weeks old, weighing approximately 180 g, purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. *Streptococcus salivarius* CCFM1512, OJSWX19B1, K12, *Fusobacterium nucleatum* (… Fusobacterium nucleatumATCC25586 and Porphyromonas gingivalis ATCC33277 were obtained from the Microbial Culture Collection Center of the Biotechnology Center of Jiangnan University. Human gingival fibroblasts (HGF) were obtained from the Cell Collection Center of the Biotechnology Center of Jiangnan University.
[0027] The culture media involved in the following examples are as follows: MRS liquid culture medium ( / L): peptone 10.0 g, beef extract 10.0 g, yeast powder 5.0 g, glucose 20.0 g, anhydrous sodium acetate 2.0 g, magnesium sulfate heptahydrate 0.5 g, manganese sulfate monohydrate 0.25 g, diammonium hydrogen citrate 2.0 g, dipotassium hydrogen phosphate trihydrate 2.6 g, Tween-80 1.0 mL.
[0028] MRS solid medium ( / L): peptone 10.0 g, beef extract 10.0 g, yeast extract 5.0 g, glucose 20.0 g, anhydrous sodium acetate 2.0 g, magnesium sulfate heptahydrate 0.5 g, manganese sulfate monohydrate 0.25 g, diammonium hydrogen citrate 2.0 g, dipotassium hydrogen phosphate trihydrate 2.6 g, Tween-80 1.0 mL, agar 18.0 g.
[0029] 0.5% heme chloride-vitamin K1 solution ( / L): 0.5 g heme chloride, 10.0 mL 1 mol / L sodium hydroxide, 10.0 mL 10 mg / mL vitamin K1 solution.
[0030] BHI liquid culture medium ( / L): 17.5 g beef heart extract, 10.0 g tryptone, 5.0 g sodium chloride, 5.0 g yeast extract, 2.5 g disodium hydrogen phosphate dodecahydrate, 2.0 g glucose, 1.0 g cysteine, 1.0 mL 0.5% vitamin K1-heme chloride.
[0031] BHI solid medium ( / L): 17.5 g bovine heart extract, 10.0 g tryptone, 5.0 g sodium chloride, 5.0 g yeast extract, 2.5 g disodium hydrogen phosphate dodecahydrate, 2.0 g glucose, 1.0 g cysteine, 1.0 mL 0.5% vitamin K1-heme chloride, 18.0 g agar, and 50 mL sterile defibrinated sheep blood.
[0032] Complete cell culture medium: 89% (v / v) DMEM medium, 10% (v / v) fetal bovine serum, 1% (v / v) 100× penicillin and streptomycin mixed solution (penicillin content 10000 U / mL, streptomycin concentration 10 mg / mL).
[0033] The methods for preparing bacterial suspension, fermentation supernatant, and bacterial lysate involved in the following examples are as follows: P. gingivalis and F. nucleatum Bacterial suspension: (1) Activation and culture of strains: All strains to be activated were stored in a -80℃ refrigerator with 30% glycerol. During activation, a sterile inoculation loop was used to dip the bacterial solution into the BHI solid medium and streak it onto the medium. The medium was then placed in an anaerobic workstation and cultured at 37℃ for 72 h. After that, a single colony was picked up with an inoculation loop and placed into the BHI liquid medium. The medium was then placed in an anaerobic workstation and cultured at 37℃ for another 48 h.
[0034] (2) The activated bacterial suspension was inoculated into BHI liquid medium at an inoculation rate of 4% (v / v) and incubated at 37°C for 36 h in an anaerobic workstation. After colony counting, the suspension was concentrated by centrifugation to adjust the bacterial suspension to 1×10⁻⁶. 9 CFU / mL.
[0035] Supernatant from Streptococcus salivarius fermentation: (1) Activation and culture of strains: All strains to be activated were stored in a -80°C refrigerator with 30% glycerol. During activation, a sterile inoculation loop was used to dip the bacterial solution into the MRS solid medium and streak it onto the medium. After culturing at 37°C for 48 h, a single colony was picked up with an inoculation loop and transferred to the MRS liquid medium. The medium was then cultured at 37°C for another 24 h.
[0036] (2) The activated bacterial culture was inoculated into MRS liquid medium at an inoculation rate of 2% (v / v), and cultured in a fermenter at 37°C for 18 h. The supernatant was collected by centrifugation and then concentrated to 5×10⁻⁶ by rotary evaporation. 9 The fermentation supernatant corresponding to a CFU / mL cell concentration is cell-free supernatant (CFS).
[0037] Streptococcus salivarius cell lysate: (1) Activation and culture of the strain: Same as above (2) The activated bacterial solution was inoculated into MRS liquid medium at an inoculation rate of 2% (v / v), and placed in a fermenter for 37°C for 18 h. The bacterial sludge was collected by centrifugation and the concentration was adjusted to 5×10⁻⁶ with sterile physiological saline. 9 CFU / mL. The bacterial lysate was then homogenized 10 times at 1200 MPa using a high-pressure homogenizer to obtain the lysate.
[0038] Example 1: Screening and identification of Streptococcus salivarius CCFM1512 Strain screening: 0.2 mL of non-irritating saliva sample collected from healthy individuals was added to 1.8 mL of sterile saline solution and mixed thoroughly to obtain strain 10. -1 The diluted solution, then take 10 -1 Add 0.5 mL of the diluent to 4.5 mL of sterile physiological saline to obtain 10. -2The diluted solution was used to obtain a series of dilution gradients following the above procedure. Take 10... -4 10 -5 10 -6 0.1 mL of each diluent was applied to the surface of the solidified MRS plate and spread with a spreader until the plate surface was basically dry. Then the plate was incubated upside down at 37°C for 48 h.
[0039] Strain identification: Single colonies were picked from the surface of MRS solid plates using an inoculation loop and incubated in MRS liquid medium at 37°C for 24 h. 1 mL of bacterial suspension was centrifuged at 6000 rpm for 3 min at 4°C. The supernatant was discarded, and the suspension was resuspended in 1 mL of sterile deionized water. The suspension was then centrifuged again at 6000 rpm for 3 min at 4°C, and the supernatant was discarded. This washing process was repeated twice, and the suspension was resuspended in 1 mL of sterile deionized water as a template for strain identification. A 25 μL PCR system was prepared, including 0.5 μL of forward primer (10 μM), 0.5 μL of reverse primer (10 μM), and 12.5 μL of 2×... Taq PCR Master Mix, 1 μL bacterial suspension, 10.5 μL double-distilled water. Primer information is shown in Table 1.
[0040] Table 1 Primer Information Table
[0041] The sequencing results were used for species confirmation using the BLAST function of the NCBI database (http: / / www.ncbi.nlm.nih.gov / ). The comparison results showed that the strain was *Streptococcus salivarius*, named CCFM1512.
[0042] Example 2: Intervention of Salicylic Streptococcus CCFM1512 supernatant to alleviate HGF inflammatory response Human gingival fibroblasts (HGF) were resuscitated, passaged three times, and then injected with 4 × 10⁻⁶ cells. 5 Cells were inoculated at a concentration of 1 cell / mL, with 2 mL of the solution added to each well of a 6-well cell culture plate. After pre-culturing at 37°C in 5% CO2 for 16 h, the culture medium was replaced with serum-free medium and incubated overnight. Except for the control group, each well was treated with Pg-LPS at a final concentration of 5 μg / mL to induce inflammation. After 2 h of stimulation, cell-free supernatant (CFS) of Streptococcus salivarius was added, and co-incubation continued for 24 h. The cell culture supernatant was then collected and pretreated according to the ELISA kit instructions, and the levels of TNF-α and PGE2 were measured. Three replicates were set up for each group.
[0043] The measurement results are as follows Figure 1As shown, under Pg-LPS stimulation, the levels of TNF-α and PGE2 in the cell supernatant of the model group were significantly higher than those in the control group, reaching 168.48 pg / mL and 259.71 pg / mL, respectively. After intervention with the supernatant of *Streptococcus salivarius* CCFM1512, the levels of TNF-α and PGE2 in the cell supernatant were significantly lower than those in the model group. p <0.001), while intervention with the fermentation supernatant of another Streptococcus salivarius strain, OJSWX19B1, could not suppress this inflammatory response ( p >0.05). Therefore, we conclude that, at the cellular level, the fermentation supernatant of Streptococcus salivarius CCFM1512 can reduce the levels of TNF-α and PGE2 in fibroblast culture supernatant and inhibit the inflammatory response.
[0044] Example 3: Application of Salicylic Streptococcus CCFM1512 supernatant in regulating inflammatory response and alleviating periodontitis in rats 1. Preparation of fermentation supernatant of Streptococcus salivarius Using a sterile inoculation loop, streak *Streptococcus salivarius* CCFM1512 culture onto MRS solid medium and incubate at 37°C for 48 h. Then, use an inoculation loop to pick single colonies and transfer them to MRS liquid medium, incubating at 37°C for another 24 h. Inoculate the activated culture at a 2% (v / v) inoculation rate into MRS liquid medium and incubate in a fermenter at 37°C for 18 h. Collect the supernatant by centrifugation and then concentrate it to 5 × 10⁻⁵ ml by rotary evaporation. 9 Fermentation supernatant corresponding to a CFU / mL cell concentration.
[0045] 2. Experimental Design SPF-grade male Wistar rats, 6 weeks old, weighing approximately 180 g, were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. *Streptococcus salivarius* CCFM1512 and K12, *Fusobacterium nucleatum* ATCC25586, and *Porphyromonas gingivalis* ATCC33277 were obtained from the Microbial Culture Collection Center of the Biotechnology Center of Jiangnan University. Figure 2 Table A shows the animal experiment procedure, and Table 2 shows the animal grouping.
[0046] Table 2 Animal grouping and diet and water intake
[0047] Before the experiment, all animals were randomly divided into four groups and placed in an environment with alternating light and dark conditions at 22-24℃ for one week to acclimatize. The control group rats were given normal food and water throughout the experiment, while the model group and experimental group rats were given a high-sugar diet (Keyes 2000) and distilled water containing 10% sucrose after the acclimatization period. The specific steps of the experiment are as follows: (1) Silk thread ligation for modeling: Days 1-7 Except for the control group, rats in the experimental group underwent silk ligation modeling on day 0. The specific procedure was as follows: Rats in the experimental group were intraperitoneally injected with an anesthetic of ketamine:toluidine = 100:10 mg / kg to induce general anesthesia. Orthodontic ligature wires with a diameter of 0.22 mm were used to ligate the left and right maxillary second molars. Excess ligature wire was buried in the gingival sulcus as much as possible to avoid affecting normal chewing. Figure 2 B). After surgery, the rats were placed in a warm and comfortable environment for half an hour to await recovery, followed by a one-week recovery period. During the recovery period, the rats were given distilled water containing 125 μg / mL ampicillin sodium for three consecutive days.
[0048] (2) Pathogen infection: Days 8-14 Blank control group: 0.5 mL of sterile PBS buffer was drawn with a 1 mL sterile syringe and slowly rinsed the rat's maxilla for 2 min, followed by fasting and water restriction for half an hour.
[0049] Experimental group: 1×10⁻⁶ liters were drawn using a 1 mL sterile syringe. 9 CFU / mL Porphyromonas gingivalis and 1×10 9 A mixture of 0.5 mL of CFU / mL Fusobacterium nucleatum was slowly rinsed at the ligation site of the rat's maxilla for 1 min, followed by fasting and water restriction for half an hour. Infection was repeated every other day for a total of four times.
[0050] (3) Intervention experiment: Days 15-42 Blank control group: 1 mL of sterile PBS buffer was drawn with a sterile syringe and slowly rinsed in the maxilla of rats for 2 min. Then, the rats were fasted and deprived of water for half an hour. This was done once a day.
[0051] Model group: 1 mL of sterile PBS buffer was drawn into a sterile syringe and slowly rinsed at the ligation site of the rat's maxilla for 2 min. Then, the rat was fasted and deprived of water for half an hour, once a day.
[0052] Streptococcus salivarius K12 cell group: 1 mL of Streptococcus salivarius K12 cell lysate was drawn with a 1 mL sterile syringe and slowly rinsed at the ligation site of the rat's maxilla for 2 min. Then, the rats were fasted and deprived of water for half an hour. This was done once a day.
[0053] Salivarius CCFM1512 supernatant group: 1 mL of Salivarius CCFM1512 fermentation supernatant was drawn with a 1 mL sterile syringe and slowly rinsed at the ligation site of the rat's maxilla for 2 min. Then, the rat was fasted and deprived of water for half an hour, once a day.
[0054] After the intervention, periodontal tissues of the rats were sampled, and the rat maxilla was separated for subsequent Micro-CT imaging, HE, Masson's Law, TRAP staining, immunofluorescence staining, and immunohistochemical staining. A shallow incision was made in the gingival tissue near the second maxillary molar of the rat (to avoid damage to the maxilla), and gingival tissue was collected with forceps for subsequent inflammatory factor measurement and qPCR experiments.
[0055] 3. Experimental Results (1) Alleviating alveolar bone resorption in rats The rat maxilla was isolated, and Micro-CT was used to scan and image it in three dimensions. Alveolar bone loss (ABL) was quantified by the distance from the cementoenamel junction (CEJ) to the alveolar ridge crest (ABC). The relevant results are as follows: Figure 3 As shown. The transverse CT cross-section and three-dimensional imaging of the second molar (…). Figure 3 A) As can be seen, compared with the control group, the alveolar bone of the model group rats showed significant resorption in both the horizontal and vertical directions, with more exposed tooth roots and wider interdental gaps. The alveolar bone gap (ABL) of the model group was 0.927 mm, significantly higher than that of the control group (0.518 mm). p <0.001), and the construction of the periodontitis model also significantly reduced the bone mineral density of the bone tissue in the vicinity of the ligation ( Figure 3 C). After intervention with the supernatant of *Streptococcus salivarius* CCFM1512, the alveolar bone resorption (ABL) in the intervention group decreased to 0.804 mm, while intervention with the positive control strain *Streptococcus salivarius* K12 failed to alleviate alveolar bone resorption. Figure 3 B).
[0056] (2) Histopathological analysis of periodontal tissue in rats Histopathological sections of the rat maxilla along the sagittal plane were stained with hematoxylin and eosin (HE). This revealed that, compared to the control group, the model group had a thinner junctional epithelium and was more distant from the CEJ attachment. Furthermore, the model group also exhibited greater bone resorption and inflammatory cell infiltration. Figure 4 A). Masson staining revealed that inflammation caused degeneration and degradation of collagen fibers around ABC in model group A, resulting in a disordered and sparse arrangement of collagen fibers in the periodontal tissues. Figure 4 B). Four weeks after intervention with the supernatant of *Streptococcus salivarius* CCFM1512, the gingival epithelial structure of rats was relatively intact compared to the model group, with denser and more regularly arranged collagen fibers, more closely resembling normal tissue. Similarly, intervention with *Streptococcus salivarius* K12 could also improve the above conditions to some extent.
[0057] (3) Expression of DEL-1 in rat periodontal tissues Immunohistochemical staining of periodontal tissues showed that, compared with the model group, intervention with CCFM1512 supernatant restored the level of DEL-1, an endothelial developmental site, in periodontal tissues. Figure 4 C). This was validated in rat gingival tissue, where intervention with CCFM1512 fermentation supernatant significantly upregulated the DEL-1 gene. EDIL3 The mRNA level was restored to a level comparable to that of the control group. Figure 4 D). Current research indicates that in the context of periodontitis, DEL-1 and IL-17 not only mediate opposite functions but also have a synergistic regulatory relationship. IL-17 directly inhibits the expression of DEL-1 in endothelial cells, thereby promoting LFA-1-dependent neutrophil recruitment. Measurements of IL-17A and the neutrophil effector marker MPO levels in rat gingival tissue showed that, compared to the model group, intervention with CCFM1512 supernatant significantly reduced the level of IL-17A in gingival tissue (D). p <0.05, which may explain the upregulation of DEL-1 ( Figure 4 E). Simultaneously, intervention with CCFM1512 supernatant reduced the MPO level in gingival tissue from 2.606 pg / mg in the model group to 1.905 pg / mg (E). Figure 4 F).
[0058] (4) Effects of M1 / M2 polarization of macrophages in rat periodontal tissues The presence of M1 macrophages (labeled with iNOS) and M2 macrophages (labeled with CD206) in periodontal tissues was analyzed by immunofluorescence staining. Figure 5 A). Compared with the control group, the model group had more M1 macrophages at the root of the second molar. In contrast, the M1 macrophages in the CCFM1512 supernatant intervention group were significantly reduced, while the M2 macrophages were increased. Similarly, we measured the levels of typical pro-inflammatory factors IL-1β and IL-6 secreted by M1 macrophages in rat gingival tissue. The results showed that the levels of IL-1β and IL-6 in the gingival tissue of the CCFM1512 supernatant intervention group were significantly lower than those in the model group. Figure 5 (B, C). Simultaneously, we also examined the genetic components of gingival tissue. iNOS (M1 macrophage markers) and Arg-1 The expression level of (M2 macrophage marker) was correlated with that in the gingival tissue of the CCFM1512 supernatant intervention group. iNOS The mRNA level was significantly lower in the group compared to the model group. Arg-1 Levels significantly increased ( Figure 5(D, E). While intervention with Salicylic Streptococcus K12 could reduce the levels of inflammatory factors in gingival tissue, it failed to significantly affect macrophage polarization.
[0059] Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Anyone skilled in the art can make various modifications and alterations without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be determined by the claims.
Claims
1. Streptococcus salivarius ( Streptococcus salivarius CCFM1512 was deposited at the Guangdong Provincial Center for Microbial Culture Collection on August 21, 2025, with accession number GDMCC No: 66863.
2. A microbial preparation containing the Streptococcus salivarius CCFM1512 as described in claim 1.
3. The metabiotic prepared from *Streptococcus salivarius* CCFM1512 as described in claim 1, characterized in that, The postgenetic agent is prepared by any of the following methods: (a) After fermenting the *Streptococcus salivarius* CCFM1512, collect the fermentation supernatant; (b) After fermenting the said Streptococcus salivarius CCFM1512, collect the bacterial cells and lyse them to obtain the bacteria.
4. A product containing the *Streptococcus salivarius* CCFM1512 of claim 1, the microbial preparation of claim 2, or the postbiotic of claim 3.
5. The product as described in claim 4, characterized in that, The products include food, health products, medicines, or daily chemical products.
6. The product as described in claim 5, characterized in that, The drug also contains conventional excipients and / or pharmaceutical carriers.
7. The product as described in claim 6, characterized in that, The conventional excipients include one or more of the following: fillers, flavoring agents, binders, disintegrants, lubricants, antacids, and nutritional fortifiers.
8. The product as described in claim 7, characterized in that, The pharmaceutical carrier includes one or more of the following commonly used medical fillers, adhesives, wetting agents, disintegrants, lubricants, and flavoring agents.
9. The product as described in claim 8, characterized in that, The daily chemical products mentioned include toothpaste, mouthwash, or oral spray.
10. The use of the *Streptococcus salivarius* CCFM1512 of claim 1 or the metabiotic of claim 3 in the preparation of a medicament for relieving oral inflammation.