JP5827704B2 - Periodontal disease marker - Google Patents

Description

  The present invention relates to a periodontal disease marker.

  Periodontal tissue refers to tissue surrounding the teeth and supporting the teeth, and is composed of gingiva, periodontal ligament, cementum, and alveolar bone. Among the inflammation of the periodontal tissue, the one where the inflammation is confined to the gingiva is called “gingivitis”, and the case where the inflamed part extends beyond the gum and the periodontal ligament and alveolar bone are damaged or destroyed is called “periodontitis” . These are collectively called “periodontal disease”.

  Gingivitis and periodontitis of periodontitis are both affected by the growth of bacteria in plaque (the mass that grew by attaching bacteria etc. to food residues in the oral cavity) Endogenous infection caused by inflammation of periodontal tissue. Periodontal disease, unlike caries, is almost painless and often progresses without being noticed. However, if the periodontal disease is left untreated, the symptom progresses, and there is a great risk that the tooth will eventually fall out, and there is also a concern that the risk of circulatory disease increases.

Periodontal disease measures include preventive actions such as correct tooth brushing, plaque control by regular checkups, and improvement of dietary habits. However, in reality, it is very important how to accurately detect a periodontal disease that is progressing by periodontal disease diagnosis and to perform treatment suitable for the symptom.
Diagnosis of periodontal disease is generally performed by measuring periodontal pockets, attachment level, X-ray image diagnosis, and the like. However, these diagnostic methods place a heavy burden on the subject. Further, these periodontal disease diagnosis methods have a complicated operation procedure and have problems such as individual differences in judgment criteria because they are based on experience and skills of dentists.

  As a method for diagnosing periodontal disease using a periodontal disease marker, gingival crevicular fluid is collected with a brush-like sampling device, and biomarkers such as lactoferrin, α1-antitrypsin, and hemoglobin contained in the gingival crevicular fluid are detected. And the method of diagnosing periodontal disease is known (for example, refer patent documents 1 and 2). However, the gingival crevicular fluid contains components other than the gingival crevicular fluid, and accuracy may be lost when saliva is mixed. Furthermore, a method of diagnosing periodontal disease by detecting occult blood (hemoglobin) in saliva or mouthwash discharge liquid is also known (see Non-Patent Document 1, for example). However, occult blood in saliva may contain other substances than those derived from periodontal disease. further. Saliva volume varies greatly between individuals and diurnal variation, and since there is no hemoglobin quantitativeness, it is impossible to diagnose the degree of progression of periodontal disease.

Japanese Patent Laid-Open No. 10-2899 JP 2000-28608 A Koichi Ito, Journal of Japanese Society of Dentistry, 24, 112-115, 2005

This invention makes it a subject to provide the marker for periodontal disease onset determination which can determine the presence or absence of onset of periodontal disease simply and correctly.
Another object of the present invention is to provide a marker for determining the degree of progression of periodontal disease that can easily and accurately determine the degree of progression of periodontal disease.
Moreover, this invention makes it a subject to provide the periodontal disease onset determination method which can determine the presence or absence of onset of a periodontal disease simply and correctly.
Moreover, this invention makes it a subject to provide the periodontal disease progress determination method which can determine the extent of a periodontal disease progress simply and correctly.
Furthermore, the present invention provides a screening method for a periodontal disease ameliorating agent or preventive agent that is effective for simple and accurate screening of a periodontal disease ameliorating agent or prophylactic agent effective in preventing and improving periodontal disease. Let it be an issue.

  Bacteria have acquired a mechanism for sensitively sensing changes in their environment. As one of such mechanisms, it is clear that it senses its own density in the environment through a specific information transmission substance and skillfully controls its various biological activities according to the density. ing. Such an information transmission mechanism between bacteria is called a quorum sensing system.

  Bacteria with a quorum sensing system synthesize and release a signaling molecule called an autoinducer and respond to the signaling molecule to control gene expression as a function of bacterial density. To date, it has been reported that autoinducer-2 is used for communication between different bacterial species (see, for example, Bassler et al., Bacteriol. 179, pp. 4043-4045, 1997).

There are about 600 species of bacteria as oral flora, these, poly Philo MONAS (Porphyromonas) bacteria, Streptococcus (Streptococcus) bacteria, Eikenera (Eikenella) bacteria, Lactobacillus (Lactobacillus) bacteria, Actinobacillus (Actinobacillus) bacteria, Actinomyces (Actinomyces) bacteria, Fusobacterium (Fusobacterium) bacteria, Beironera (Veillonella) bacteria, capsules Roh site file moth (Capnocytophaga) bacteria and Prevotella (Prevotella) bacteria such as multiple teeth It is known that peripathogenic bacteria also exist. When these periodontal pathogens proliferate, secrete exopolysaccharides, and form a biofilm, pathogenicity increases and periodontal disease develops.
Japanese Patent Application Laid-Open No. 2008-214296 discusses the biofilm formation inhibitory effect when an antagonist of an autoinducer-2 receptor is added using a Porphyromonas gingivalis strain, which is one of periodontal pathogens. However, there are many other bacterial species other than Porphyromonas gingivalis . Moreover, there exists an effect | action with respect to periodontal pathogens of oral resident bacteria other than periodontal pathogens. Therefore, even if the biofilm formation inhibitory effect is examined using only the Porphyromonas gingivalis strain, the level and severity of periodontal disease in mammals such as humans cannot be clinically explained.

In view of the above problems, the present inventors have conducted intensive studies. As a result, the influence of autoinducer-2 on periodontal disease in vivo is clarified, and autoinducer-2 used for communication between different bacterial species released when resident bacteria present in the oral cavity grow. It was found that there is a correlation between the amount of cancer and periodontal disease. Specifically, it was found that the presence or absence of periodontal disease and the degree of progression of periodontal disease are related to the amount of autoinducer-2 in the oral cavity. Furthermore, it discovered that the substance which has an autoinducer-2 activity inhibitory effect was effective in periodontal disease improvement and prevention. The present invention has been completed based on these findings.

The present invention relates to a periodontal disease onset marker comprising autoinducer-2.
The present invention also relates to a marker for determining the degree of progression of periodontal disease comprising autoinducer-2.
In addition, the present invention collects a sample from the oral cavity of a subject, quantifies autoinducer-2 contained in the collected sample, and periodontal disease develops when the amount of quantified autoinducer-2 is large. The present invention relates to a method for determining periodontal disease onset.
Further, the present invention collects a sample from the oral cavity of a subject, quantifies autoinducer-2 contained in the collected sample, and periodontal disease progresses as the amount of quantified autoinducer-2 increases. The present invention relates to a periodontal disease progress determination method.
Furthermore, this invention mixes the bacterium which has a quorum sensing system via an autoinducer-2, and a test agent, and uses the test agent which has an autoinducer-2 activity inhibitory effect as a periodontal disease improving agent or a preventive agent. The present invention relates to a screening method for a periodontal disease improving agent or a preventive agent to be selected.

According to the marker for the onset of periodontal disease of the present invention, the presence or absence of the onset of periodontal disease can be determined easily and accurately.
Moreover, according to the marker for periodontal disease progression degree determination of the present invention, the degree of progression of periodontal disease can be easily and accurately determined.
Moreover, according to the method for determining the onset of periodontal disease of the present invention, the presence or absence of the onset of periodontal disease can be determined easily and accurately.
Moreover, according to the periodontal disease progression degree determination method of the present invention, the degree of progression of periodontal disease can be easily and accurately determined.
Furthermore, the method for screening for a periodontal disease improving agent or preventive agent of the present invention can conveniently and accurately screen for a periodontal disease improving agent or prophylactic agent effective in preventing and improving periodontal disease.

1 (a) to 1 (c) show, in Example 2, a control (FIG. 1 (a)), 4-bromo-5-5 as a test agent on the lower first molar (circled part) of a Syrian hamster. (4-methoxyphenyl) -2 (5H) -furanone (FIG. 1 (b)) and 3,4-dibromo-5-hydroxy-2 (5H) -furanone (FIG. 1 (c)) were respectively administered; It is a drawing-substituting photograph taken under a stereomicroscope after extracting the mandible after 2 weeks.

Hereinafter, the present invention will be described in detail based on preferred embodiments thereof.
The marker for determining the onset of periodontal disease of the present invention (also simply referred to as a marker for determining periodontal disease) and the marker for determining the progress of periodontal disease are from autoinducer-2 collected from the oral cavity of a subject such as a human. Become. Autoinducer-2 (hereinafter also referred to as “AI-2”) in the present invention is not particularly limited as long as it is produced by bacteria and has AI-2 activity.

  In the present invention, AI-2 activity refers to an activity in which AI-2 affects bacteria having a quorum sensing system, that is, an activity that promotes the function of bacteria brought about by quorum sensing via AI-2. Bacteria are luminescent, swarming, biofilm formation, proteolytic enzyme production, antibiotic synthesis, gene acceptability development, plasmid conjugation transmission, virulence factor production and spore formation by quorum sensing via AI-2 Etc. are known to perform. Thus, AI-2 activity is, in other words, bioluminescence, swarming, biofilm formation, production of proteolytic enzymes, antibiotics by bacteria that recognize AI-2, ie bacteria having the AI-2 receptor. It can be referred to as activities such as synthesis, development of gene acceptability, plasmid conjugation transmission, pathogenic factor production and sporulation. Examples of the pathogenic factor include enterotoxin, adenylate cyclase toxin, adhesin, alkaline protease, hemolytic toxin, anthrax toxin, APX toxin, α toxin, β toxin, δ toxin, C2 toxin, C3 toxin, botulinum toxin, bundled line Hair structure subunit, C5A peptidase, cardiotoxin, chemotaxis, cholera toxin, ciliary toxin, clostridial cytotoxin, clostridial neurotoxin, collagen adhesion gene, cytolysin, vomiting toxin, endotoxin, exfoliating toxin, external Toxin, extracellular elastase, fibrinogen, fibronectin binding protein, fibrillar hemagglutinin, fimbria, gelatinase, hemagglutinin, leukotoxin, lipoprotein signal peptidase, listeriolysin O, M protein, neurotoxin, non-fimbria adhesin Edema factor, Examples include, but are not limited to, hyperenzymes, pertussis toxin, phospholipase, pili, pore forming toxin, proline permease, serine protease, Shiga toxin, tetanus toxin, thiol activated cytolysin, tracheal cell lysin, urease .

  Specific examples of AI-2 in the present invention include 4,5-dihydroxy-2,3-pentanedione (DPD) represented by the following formula.

When DPD binds to the bacterial AI-2 receptor, it incorporates boron and is converted to furanosyl borate diester. Another specific example of AI-2 in the present invention is the furanosyl borate diester.
Specific examples of the furanosyl borate diester are shown below. However, the present invention is not limited to these.

The bacterium having an AI-2 receptor, e.g., Porphyromonas (Porphyromonas) bacteria, Haemophilus (Haemophilus) bacteria, Bacillus (Bacillus) bacteria, Neisseria (Neisseria) bacteria, Streptococcus (Streptococcus) bacteria, Eikenera (Eikenella) bacteria belonging to the genus Lactobacillus (Lactobacillus) bacteria belonging to the genus Actinobacillus (Actinobacillus) bacteria belonging to the genus Actinomyces (Actinomyces) bacteria belonging to the genus Bacteroides (Bacteroides) bacteria belonging to the genus, Cap Roh site file moth (Capnocytophaga) bacteria belonging to the genus, Fusobacterium (Fusobacterium) bacteria, Peputokokkasu (Peptococcus) bacteria, Prevotella (Prevotella) bacteria, Seremonasu (Selemonas) bacteria, such as Eubacterium (Eubacterium) bacteria and Beironera (Veillonella) bacteria and the like. More specifically, poly Philo Monas gingivalis (Porphyromonas gingivalis), Streptococcus pyogenes (Streptococcus pyogenes), Streptococcus mutans (Streptococcus mutans), Eikenera-Korodensu (Eikenella corrodens), Lactobacillus salivarius (Lactobacillus salivarius), Streptococcus Sanginisu (Streptococcus sanguinis), Streptococcus anginosus (Streptococcus anginosus), Streptococcus oralis (Streptococcus oralis), Streptococcus Gorudoni (Streptococcus gordonii), Streptococcus mitis (Streptococcus mitis), Actinobacillus actinomycetemcomitans (Actinobacillus actinomycetemcomitans), Lactobacillus rhamnosus (Lactobacillus rhamnosus), Actinomyces Naesurandi Actinomyces naeslundii), Fusobacterium nucleatum (Fusobacterium nucleatum), Beironera-Parla (Veillonella parvula), Cap Roh site file moth-Suputigena (Capnocytophaga sputigena) and Prevotella interface media (Prevotella intermedia) and the like.

  In the present invention, AI-2 is collected from the oral cavity of a subject such as a human and used. Here, examples of the subject include mammals such as humans, monkeys, chimpanzees, dogs, cats, cows, pigs, rats, and mice. The AI-2 is not particularly limited as long as it is collected from the oral cavity, and AI-2 contained in specimens such as saliva, plaque, tongue coating, and gingival crevicular fluid collected from the oral cavity can be used. . In the present invention, it is preferable to use AI-2 contained in dental plaque. The saliva can be obtained as it is discharged as it is or whether it is impregnated with water, regardless of whether it is unstimulated or stimulated. Plaque can be collected by cleaning the tooth surface with a dental scaler, swab, brush, or inserting a paper point. Tongue moss is obtained by brushing with a brush, cotton swab or gauze. The gingival crevicular fluid can be obtained by inserting a paper point or the like into the gingival sulcus. However, the method for collecting AI-2 is not limited to this.

  The present inventors have found that when the amount of AI-2 present in the oral cavity of the subject is large, the subject has developed periodontal disease. Furthermore, it discovered that periodontal disease progressed, so that there were many amounts of AI-2. Therefore, by quantifying the amount of AI-2 collected from the oral cavity, it is possible to determine the presence or absence of the onset of periodontal disease and the degree of progression (severity) of the periodontal disease. As an example, when the AI-2 activity in the case of 10 μM DPD is 100, when the AI-2 activity is 0 or more and less than 3, normal / mild periodontal disease, and when it is 3 or more and less than 10, moderate periodontal disease, 10 The above can be determined to be severe periodontal disease. Further, the therapeutic effect can be determined by comparing the amount of AI-2 before the start of periodontal disease treatment and the amount of AI-2 after treatment.

There is no restriction | limiting in particular as a quantification method of AI-2, It can carry out by a normal method (For example, Chen X. et al., Nature, vol. 415, p. 545-549, 2002; and Thiel V. et. al., Chembiochem., vol. 10 (3), p. 479-485, 2008).
For example, AI-2 can be quantified by a bioassay that measures luminescence intensity using a reporter bacterium that emits light by recognizing AI-2, preferably a bacterium having AI-2 receptor and luciferase activity. it can. More specifically, Vibrio Harvey BB170 strain is used as an AI-2 reporter bacterium, and 4-hydroxy-5-methyl-3 (2H) -furanone (HMF) standard and / or DPD standard as a standard sample for AI-2. A calibration curve can be prepared and AI-2 can be quantified from the luminescence intensity when a test sample is added (for example, Chen X. et al., Nature, vol. 415, p. 545-549, 2002). reference). Moreover, after reacting AI-2 contained in a test sample with phenylenediamine and N-methyl-N- (trimethylsilyl) trifluoro-acetamide, AI-2 can be quantified by GC-MS (for example, Thiel V. et al., Chembiochem., Vol. 10 (3), p. 479-485, 2008). However, the present invention is not limited to these.
A test sample for quantifying AI-2 is preferably subjected to ultrafiltration of a specimen collected from the oral cavity with a filter having a cutoff molecular weight of 3000 or less after the crushing / extraction process to remove impurities.
In the present invention, it is preferable to use a container having low AI-2 adsorptivity, such as a glass container, for preparing and storing a test sample. By using a glass container, AI-2 contained in the test sample can be accurately quantified.

In the screening method for periodontal disease ameliorating agent or preventive agent of the present invention, a test agent having an autoinducer-2 activity inhibitory effect by mixing a bacterium having a quorum sensing system via autoinducer-2 with a test agent Is selected as a periodontal disease improving agent or preventive agent. The bacterium having a quorum sensing system via autoinducer-2 is not particularly limited, and a reporter bacterium that recognizes autoinducer-2 and emits light is preferable, and oral bacteria and periodontal pathogens can also be used. In addition, bacteria having a quorum sensing system via autoinducer-2 collected from the oral cavity of the subject can also be used as a specimen.
The “autoinducer-2 activity inhibitory effect” in the present invention refers to an inhibitory effect on AI-2 activity. Specifically, bioluminescence, swarming, biofilm formation, production of proteolytic enzymes, synthesis of antibiotics, development of gene acceptability, plasmid conjugation transmission, pathogenesis by the aforementioned bacteria having AI-2 receptor It refers to inhibitory effects on AI-2 activity such as factor production and sporulation.

  EXAMPLES Hereinafter, although this invention is demonstrated further in detail based on an Example, this invention is not limited to this.

Example 1 Determination of Periodontal Disease Onset and Periodontal Disease Progression (1) Determination of AI-2 in plaque Vibrio Harvey BB170 strain (distributed from ATCC, AB (Autoinducer Bioassay) medium (10 mM potassium phosphate buffer [pH 7] 0.0], 0.3 M NaCl, 0.05 M MgSO ₄ , 0.2% casamino acid, 2% glycerol, 1 mM L-arginine, 1 μg / mL thiamine, 0.01 μg / mL riboflavin) at 30 ° C. overnight. Culture) was used as AI-2 reporter bacteria and diluted 5000 times in AB medium to prepare a reporter bacteria solution. Various concentrations of HMF preparation (manufactured by Sigma) or DPD preparation (manufactured by OMM Scientific) were mixed with the reporter bacterial solution so that the volume ratio of the reporter bacterial solution: preparation was 9: 1, and the mixture was favorably mixed at 30 ° C. Air shaking culture was performed. The luminescence intensity after 4 hours was measured with a chemiluminescence meter (LB940 type, manufactured by Bertrud Japan), and a calibration curve regarding the luminescence intensity and the amount of AI-2 was prepared.

  Of the 94 male and female subjects in their 20s to 90s, subjects who collected samples in the morning stopped brushing after brushing before sleeping on the day before the test, and those in the afternoon after brushing on the morning of the test day. In addition, eating, drinking and smoking were prohibited 30 minutes before sampling.

  A total amount of 0.001 to 100 mg of subgingival plaque adhering to the tooth surface of one tooth was collected with a dental scaler. The collected plaque was dispersed in 1 mL of PBS, incubated at 90 ° C. for 10 minutes, and then crushed with 0.3 g of zirconia / silica beads (particle size: 0.1 mm, manufactured by Tommy Seiko Co., Ltd.). After centrifuging at 14,000 rpm for 5 minutes, the supernatant was passed through a 0.22 μm filter (Ultra Free MC, manufactured by Millipore) and a filter with a molecular weight of 3000 cut (Microcon YM-3, manufactured by Millipore) to prepare a sample. . The sample prepared in this manner was mixed with the reporter bacterial solution prepared in the same manner as described above at a reporter bacterial solution: standard volume ratio of 9: 1, cultured at 30 ° C. under aerobic shaking, and cultured for 4 hours. The later emission intensity was measured with a chemiluminescence meter. AI-2 contained in dental plaque was quantified from the measured luminescence intensity and the calibration curve.

(2) Relationship between the amount of AI-2 in dental plaque and the periodontal pocket depth (PD) A dental probe was inserted into the subject's periodontal pocket, and the periodontal pocket depth (PD) was measured. Rated by stage. The depth of the periodontal pocket (PD) was measured every 1 mm.
(PD)
3 mm or less: Healthy / mild periodontal disease (gingivitis)
4-5 mm: Moderate periodontal disease (periodontitis)
6 mm or more: Severe periodontal disease (periodontitis)

  Table 1 shows the relationship between the PD measured according to the above method and the amount of AI-2 quantified in (1).

  As is apparent from Table 1, there is a correlation between the amount of AI-2 in plaque and PD, and the amount of AI-2 contained in subgingival plaque increases as PD becomes deeper. Therefore, by quantifying the amount of AI-2 in the oral cavity of the subject, the progress of periodontal disease can be determined easily and accurately.

(3) Relationship between the amount of AI-2 in dental plaque and the gingival inflammation index (GI) A dental probe was inserted into the periodontal pocket of the subject, and the degree of inflammation of the gingiva was evaluated in four stages. The determination of Gingival Index (GI) score was as follows.
(GI)
0: Normal gingiva 1: Inflamed gingiva. No bleeding with probing.
2: The gums are inflamed. Probing and bleeding.
3: Spontaneous bleeding / tumor formation

  Table 2 shows the relationship between the GI measured according to the above method and the amount of AI-2 quantified in (1).

  As is clear from Table 2, there is a correlation between the amount of AI-2 in plaque and GI, and when inflammation occurs in the gingiva, the amount of AI-2 contained in subgingival plaque increases. Therefore, by determining the amount of AI-2 in the oral cavity of the subject, the presence or absence of periodontal disease can be determined easily and accurately.

(4) Relationship between the amount of AI-2 in dental plaque and the bleeding index (BOP) A dental probe was inserted into the periodontal pocket of the subject, and the presence or absence of bleeding from the periodontal pocket was evaluated in four stages. Bleeding index (Bleeding on Probing, BOP) score was determined as follows.
(BOP)
0: No bleeding 1: With punctate bleeding 2: With linear bleeding 3: With natural bleeding

  Table 3 shows the relationship between BOP measured according to the above method and the amount of AI-2 quantified in (1) above.

  As is apparent from Table 3, there is a correlation between the amount of AI-2 in plaque and BOP, and when bleeding occurs in the gingiva due to probing, the amount of AI-2 contained in subgingival plaque increases. Yes. Therefore, by determining the amount of AI-2 in the oral cavity of the subject, the presence or absence of periodontal disease can be determined easily and accurately.

  From the above results, it was clinically shown that there is a correlation between the amount of AI-2 in the oral cavity and periodontal disease. Therefore, AI-2 present in the oral cavity can be used as a periodontal disease onset determination marker and a periodontal disease progression degree determination marker.

Example 2 Screening for Periodontal Disease Improvement Agent or Preventive Agent Vibrio Harvey BB170 strain (cultured overnight at 30 ° C. in AB medium) as AI-2 reporter bacteria, diluted 4500 times in AB medium, reporter bacteria A liquid was prepared. After mixing the reporter bacterial solution and each test agent and preincubating at room temperature for 10 minutes, DPD was added, aerobic shaking culture was performed at 30 ° C., and the luminescence intensity after 4 hours was measured with a chemiluminescence meter (LB940 type, belt-type). Manufactured by Ludo Japan). From the measured emission intensity, AI-2 was quantified in the same manner as in Example 1.

  As test agents, compound 1: 4-bromo-5- (4-methoxyphenyl) -2 (5H) -furanone (manufactured by Sigma), compound 2: 3,4-dibromo-5-hydroxy-2 (5H) -Furanone (manufactured by Sigma), compound 3: 4-bromo-5-methoxy-5- (4-methoxyphenyl) -2 (5H) -furanone (manufactured by Sigma), and compound 4: 4-{[3- Bromo 2- (4-methoxyphenyl) -5-oxo-2,5-dihydro-2-furanyl] oxy} benzoic acid (manufactured by Sigma) was used. As positive controls, three compounds known as AI-2 inhibitor compounds (compound 5: 4-hydroxy-2,5-dimethyl-3 (2H) -furanone (manufactured by Sigma), compound 6: 2-methoxy- 2,4-diphenyl-3 (2H) -furanone (Sigma), compound 7: 2-pentyl-2-cyclopenten-1-one (Sigma)) (Bassseler et al., Bacteriol. 179, p. 4043-4045, 1997; Yoshida A. et al., Appl. Environ. Microbiol., 71 (5), p. 2372-2380, 2005; and Wen Z. T. et al., J. Bacteriol. 189 (9), p.2682-2691, 2004).

  The amount of AI-2 was defined as a relative value to the control DPD amount (AI-2 amount) when the control agent was added with only DPD without adding the test agent. The results are shown in Table 4.

  As shown in Table 4, it was found that Compounds 1 to 4 have an AI-2 activity inhibitory effect equivalent to or higher than that of known AI-2 inhibitory compounds.

  The mandible of a Syrian hamster (7 weeks old, 3 males) was removed under anesthesia and photographed under a stereomicroscope. The evaluation site was the lingual side of the extracted mandible, and the obtained photograph was subjected to image analysis, and the vertical distance from the first molar mesial cusp to the alveolar crest was measured.

Another Syrian hamster (7 weeks old, 3 males each) was ligated with silk thread to the lower left and right first molar teeth neck of the lower jaw, and from the next day, 1% DMSO-containing PBS solution (concentration: 10 μM) of the above compound 1 or 2 Alternatively, 200 μL of PBS containing 1% DMSO was dropped twice a day for 2 weeks and reared.
After the rearing, the mandible was removed under anesthesia and photographed under a stereomicroscope. The photomicrographs are shown in FIGS. The evaluation site was the lingual side of the extracted mandible, and the obtained photograph was subjected to image analysis. The obtained photograph was used for image analysis, and the vertical distance from the first molar mesial cusp to the alveolar crest was measured.

  The alveolar bone resorption depth (value after addition for 2 weeks−value before addition) was measured from the vertical distance from the first molar mesial apex to the alveolar crest before addition of the test agent and after addition for 2 weeks. The results are shown in Table 5.

From FIG. 1 and Table 5, by dripping the compounds 1 and 2 having an AI-2 activity inhibitory effect into the oral cavity, the absorption of alveolar bone is suppressed, and the amount of AI-2 present in the oral cavity and periodontal disease It was shown that a compound having an AI-2 activity inhibitory effect is effective for the prevention and improvement (treatment) of periodontal disease.
Therefore, by selecting a test agent having an AI-2 activity inhibitory effect, it is possible to screen for a periodontal disease improving agent or preventive agent.

Claims (3)

Autoinducer-2 activity is quantified by mixing bacteria having a quorum sensing system via autoinducer-2 with a test agent that is a candidate for a periodontal disease ameliorating agent or prophylactic agent that suppresses alveolar bone resorption. Measured
The autoinducer-2 activity when the bacteria having the quorum sensing system via autoinducer-2 and 4,5-dihydroxy-2,3-pentanedione are mixed without mixing the test agent is 100. when, for this value, a test agent quantitative value of autoinducer-2 activity when mixed with the test agent is less than 29.6% as a relative value, it has a autoinducer-2 activity inhibitory effect Select as periodontal disease ameliorating agent or preventive agent to suppress alveolar bone resorption ,
A screening method for an agent for improving or preventing periodontal disease that suppresses alveolar bone resorption . Wherein the bacterium is a Vibrio harveyi BB170 strain claim 1 Symbol placement screening methods. The screening method according to claim 1 or 2 , wherein the periodontal disease is periodontitis.