Application of ardisia crenata or extract of ardisia crenata in preparation of medicine for treating periodontitis with colitis

Abstract

The invention provides application of ardisia crenata or an extract thereof in preparation of a medicine for treating periodontitis. The invention also provides application of the ardisia crenata or the extract thereof in preparation of medicines for treating ulcerative colitis. The invention further provides application of the ardisia crenata or the extract of the ardisia crenata to preparation of a medicine for treating periodontitis with colitis. The invention verifies the treatment effect of ardisia crenata or the extract thereof on periodontitis and colitis. A new thought and a new method are provided for researching the action mechanism of the traditional Chinese medicine through integrated analysis of multi-omics and network pharmacology, a foundation is provided for deeply researching the pharmacological mechanism of the ardisia crenata for treating periodontitis, and a theoretical basis is provided for clinical application of the ardisia crenata; important scientific significance and clinical value are realized on the modern research of traditional Chinese medicine compounds and the research and development of new medicines.

Description

Technical Field

[0001] The invention relates to use of cinnabari root or an extract thereof in preparing a medicine for treating periodontitis associated with colitis. Background Art

[0002] Cinnabar root is the dried root of Ardisia crenata Sims, a plant of the Myrsinaceae family. It is a special ethnic medicine in Guizhou Province and a good medicine for the throat of the Miao people. It is included in the "Chinese Pharmacopoeia" (2020 edition) and the "Quality Standards for Traditional Chinese and Ethnic Medicinal Materials in Guizhou Province" (2003 edition). Cinnabar root mainly includes a variety of chemical components such as coumarins, triterpenoid saponins, phytosterols and benzoquinones; pharmacological studies have shown that cinnabar root has anti-inflammatory, antibacterial, immunosuppressive and anti-tumor effects. It has been clinically used to treat sore throat, tonsillitis, toothache, various inflammatory and tumor diseases.

[0003] Periodontitis is a chronic multifactorial inflammatory disease with plaque biofilm as the key cause. Its main characteristics are the destruction of periodontal supporting tissues, namely clinical attachment loss, alveolar bone resorption, periodontal pocket formation, gingival bleeding, etc. Periodontitis is a common oral disease in daily life, with a total incidence of 11.2%, and approximately 743 million people will suffer from this disease. As the disease continues to progress, late-stage periodontitis will not only cause pathological migration and lead to tooth loss, but also increase the risk of obesity, diabetes, Alzheimer's disease, atherosclerosis, adverse pregnancy outcomes, rheumatoid arthritis, inflammatory bowel disease, and colon cancer in patients, seriously affecting the patient's overall health and quality of life.

[0004] Ulcerative colitis (UC) is a chronic, relapsing inflammatory bowel disease (IBD) of the colonic mucosa. Unlike Crohn's disease, UC is mainly confined to the colonic mucosa and submucosa, starting with mucosal inflammation in the rectum and extending proximally. It specifically affects the colon and rectum through multifactorial mechanisms related to genetic changes, environmental factors, microbiota, and immune system disorders. Its typical symptoms are surface mucosal ulcers, rectal bleeding, diarrhea, and abdominal pain. Evidence has shown that UC is associated with a significantly increased risk of periodontitis and worsening oral health, and that periodontitis salivary microbiota can accelerate the progression of colitis. Fusobacterium nucleatum can not only promote alveolar bone loss, but also colonize in infected colonic tissues, ultimately leading to increased expression of IL-1β and TNF-α, decreased expression of IL-10, abnormal bile acid metabolism, and impaired epithelial barrier in the colon.

[0005] Application number: CN201410330136.9, invention name: A medicine for treating ulcerative colitis, discloses a medicine for treating ulcerative colitis, which is made of the following Chinese medicinal materials in parts by weight: 20-40 parts of Cucurbita churundis seeds, 3-9 parts of Codonopsis pilosula, 3-9 parts of Rhizoma Dioscoreae, 3-9 parts of Cucumis melon seeds, 3-9 parts of Fritillaria cirrhosa, 3-9 parts of Cinnabaris root, 3-9 parts of Folium sphaerocephalae, 3-9 parts of Caraway seeds, 3-9 parts of Star anise, 3-9 parts of Chrysanthemum indica, 1-5 parts of Blueberry, 1-5 parts of Kudingcha, 1-5 parts of Coriander, 1-5 parts of Tiger Thorn root, 1-5 parts of Morus alba, 1-5 parts of Centella asiatica, and 1-5 parts of Ginkgo biloba leaves.

[0006] CN201710785012.3, invention name: Application of bergenin in preventing and treating inflammatory bowel disease, relates to the application of bergenin, the main active ingredient of bergenia, in the preparation of drugs for preventing and treating colitis. Pharmacodynamic experiments have shown that bergenin has the effect of preventing and treating colitis, especially treating ulcerative colitis and Crohn's disease.

[0007] Cha Jun, et al., Research progress of Miao medicine Cinnabar root, Science and Technology Association Forum, 2013-12-25, Fan Weifeng, et al., Research on the content of bergenin in different parts of Cinnabar root in Guangxi, Chinese Journal of Traditional Chinese Medicine Information, 2009-01-15, The HPLC published to determine the content of bergenin in the roots and stems of Cinnabar root in different regions of Guangxi, the results showed that the components of the roots and stems of the same Cinnabar root were basically the same, and the content in the stems was greater than that in the roots, and the content of bergenin in the leaves was the lowest. The specific values ​​are: the content of bergenin in the roots of Cinnabar root produced in Nanning is 1.86%, and the content in the stems is 2.32%; the content in the roots of Cinnabar root produced in Baise is 1.76%, and the content in the stems is 2.47%; the content in the roots of Cinnabar root produced in Guilin is 1.92%, and the content in the stems is 2.53%. The 2020 edition of the Chinese Pharmacopoeia stipulates that the mass fraction of bergenin in Cinnabar root decoction pieces shall not be less than 1.0%. Although bergenin is the signature ingredient of cinnabar root, its content in the plant is low and does not reach an effective medicinal dose, and cannot reflect the efficacy of the medicinal material. Summary of the invention

[0008] The technical solution of the present invention is to provide a new use of cinnabari root or its extract.

[0009] The invention provides use of cinnabari root or an extract thereof in preparing a medicine for treating periodontitis.

[0010] The invention provides use of cinnabari root or an extract thereof in preparing a medicine for treating ulcerative colitis.

[0011] The invention provides use of cinnabari root or an extract thereof in preparing a medicine for treating periodontitis associated with colitis.

[0012] Wherein, the cinnabari root extract is cinnabari root water extract.

[0013] The preparation method of the cinnabar root water extract is as follows: add water to the cinnabar root, heat and reflux to extract, combine the filtrate, filter and concentrate to 1g / mL (based on crude drug amount).

[0014] Among them, the drug has the functions of downregulating inflammatory factors, repairing damaged barriers ZO-1 and Occludin, inhibiting IL-17 signaling pathway, JAK-STAT signaling pathway, affecting PPAR signaling pathway, linoleic acid metabolism, and regulating immune response.

[0015] The research on the medicinal value of cinnabar root in existing literature mainly focuses on pharyngitis. However, there is almost no research on its role in other inflammations. The oral cavity is the starting point of the digestive tract and is closely connected with the intestines. Existing studies have also proved that there is a correlation between periodontitis and colitis. Therefore, the present invention verifies the therapeutic effect of cinnabar root or its extract in periodontitis and colitis.

[0016] Secondly, a research system for the regulation of periodontitis phenotype by cinnabar root was constructed to clarify the mechanism of action of cinnabar root in treating periodontitis from the metabolic level. Thirdly, transcriptomics and proteomics techniques were used to screen out differentially expressed genes and proteins, and the mechanism of action of cinnabar root in treating periodontitis was analyzed through multi-omics integration and molecular biological verification.

[0017] The present invention adopts the integrated analysis of multi-omics and network pharmacology to provide new ideas and methods for studying the mechanism of action of traditional Chinese medicine, provides a basis for in-depth research on the pharmacological mechanism of cinnabar root in the treatment of periodontitis, and provides a theoretical basis for the clinical application of cinnabar root. It has important scientific significance and clinical value for the modernization research of traditional Chinese medicine compound and the development of new drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 HPLC graphs of reference solution (A) and water extract of cinnabar root (B);

[0019] Figure 2 Construction of experimental animal models;

[0020] Figure 3 Flow chart of grouping and drug administration of experimental animal models;

[0021] Figure 4 Transcriptome sequencing flow chart;

[0022] Figure 5 Growth status and weight changes of mice in each group (Figure A: weight on the day of autopsy; Figure B: DAI score; Figure C, D: colon length; Note: NC: normal control group; P: periodontitis group; PU: periodontitis + colitis group; PD: periodontitis + cinnabar root group; PUD: periodontitis + colitis + cinnabar root group; data are presented in Indicates, one-way analysis of variance (n=10); ***P<0.001, **P<0.01,

[0023] *P<0.05);

[0024] Figure 6 Effects of cinnabar root on the pathological morphology of periodontal tissues in mice with periodontitis and colitis (Note: NC: normal control group; P: periodontitis group; PU: periodontitis + colitis group; PD: periodontitis + cinnabar root group; PUD: periodontitis + colitis + cinnabar root group; b: alveolar bone, e: gingival epithelium, r: tooth root);

[0025] Figure 7 Effect of cinnabar root on alveolar bone height in mice with periodontitis and colitis (Figure A: Micro-CT 3D and 2D reconstruction of maxillary bone; Figure B: CEJ-ABC; Note: NC: normal control group; P: periodontitis group; PU: periodontitis + colitis group; PD: periodontitis + cinnabar root group; PUD: periodontitis + colitis + cinnabar root group; data are presented in Indicates, one-way ANOVA (n=10); ***P<0.001, **P<0.01, *P<0.05);

[0026] Figure 8 Effects of cinnabar root on the pathological morphology of colon tissue in mice with periodontitis and colitis (Note: NC: normal control group; P: periodontitis group; PU: periodontitis + colitis group; PD: periodontitis + cinnabar root group; PUD: periodontitis + colitis + cinnabar root group);

[0027] Fig. 9 Effects of cinnabar root on the expression levels of ZO-1 and Occludin in mice with periodontitis and colitis (Note: NC: normal control group; P: periodontitis group; PU: periodontitis + colitis group; PD: periodontitis + cinnabar root group; PUD: periodontitis + colitis + cinnabar root group);

[0028] Fig.10 Effects of cinnabar root on the pathological morphology of heart, liver and kidney tissues in mice with periodontitis and colitis (Note: NC: normal control group; P: periodontitis group; PU: periodontitis + colitis group; PD: periodontitis + cinnabar root group; PUD: periodontitis + colitis + cinnabar root group);

[0029] Fig.11Effects of cinnabar root on the expression of inflammatory factors in mice with periodontitis and colitis (D: expression levels of TNF-α, IL-10, IL-1β, and IL-6 in serum of each group; EH: expression levels of TNF-α, IL-10, IL-1β, and IL-6 in colon of each group; Note: NC: normal control group; P: periodontitis group; PU: periodontitis + colitis group; PD: periodontitis + cinnabar root group; PUD: periodontitis + colitis + cinnabar root group; data are presented in Indicates, one-way ANOVA (n=10); ***P<0.001, **P<0.01, *P<0.05);

[0030] Fig.12 PLS-DA scores of fecal metabolism of mice in each group under positive ion mode (A) and negative ion mode (B);

[0031] Fig.13 OPLS-DA scores of fecal metabolism of mice in each group under positive ion mode (A) and negative ion mode (B);

[0032] Fig.14 Volcano plots of differential metabolites in fecal metabolism of mice in each group under positive ion mode (A) and negative ion mode (B);

[0033] Fig.15 Differential metabolite clustering heat map (Figure A: NC1 group and P1 group; Figure B: P1 group and PD1 group; Note: columns represent samples, rows represent metabolites, the clustering tree on the left is the differential metabolite clustering tree, and the top is the sample clustering tree. The gradient color represents the size of the quantitative value. The redder the color, the higher the expression, and the bluer the color, the lower the expression);

[0034] Fig.16 Cluster heat map of differential metabolites (Figure A: P2 group and PU2 group; Figure B: PU2 group and PUD2 group);

[0035] Fig.17 Differential metabolite association heat map (Figure A: NC1 group and P1 group; Figure B: P1 group and PD1 group; Figure C: P2 group and PU2 group; Figure D: PU2 group and PUD2);

[0036] Fig.18 KEGG pathway enrichment of differential metabolites (Figure A: NC1 group and P1 group; Figure B: P1 group and PD1 group; Figure C: P2 group and PU2 group; Figure D: PU2 group and PUD2);

[0037] Fig.19 Main component analysis chart (Panel A: NC group and PU group; Panel B: PU group and PUD group);

[0038] Fig. 20 Volcano plot of differentially expressed genes;

[0039] Fig.21 Heat map of hierarchical clustering of differentially expressed genes;

[0040] Fig. 22 GO enrichment analysis of differentially expressed genes;

[0041] Fig.23 KEGG pathway enrichment analysis of differentially expressed genes;

[0042] Fig.24 GSEA enrichment analysis (Figure A: IL-17 signaling pathway; Figure B: JAK-STAT signaling pathway);

[0043] Fig.25 The expression of IL-17A, IL-17F, JAK2 and STAT3 mRNA in the PU group and the PUD group (Figure A: IL-17A mRNA; Figure B: IL-17F mRNA; Figure C: JAK2 mRNA; Figure D: STAT3 mRNA; Note: analyzed by t test; ***P<0.001, **P<0.01, *P<0.05). DETAILED DESCRIPTION

[0044] Example 1 Preparation of Cinnabaris Root Extract of the Present Invention

[0045] Take 200g of cinnabar root and add water (10 times the amount) to perform heating reflux extraction twice, each time for 2 hours, combine the filtrates, filter and concentrate to 1g / mL (based on crude drug amount), and store at -20°C.

[0046] The beneficial effects of the present invention are demonstrated by pharmacodynamic tests as follows.

[0047] Test Example 1: Efficacy test of the extract of cinnabar root of the present invention in treating periodontitis with colitis

[0048] 1 Materials and methods

[0049] 1.1 Main experimental materials

[0050] 1.1.1 Source of Cinnabar Root Samples

[0051] The animal dosing samples were from Guiyang, Guizhou, and provided by Guizhou Sanli Pharmaceutical Co., Ltd.

[0052] 1.1.2 Experimental animals

[0053] C57BL / 6JSPF grade male mice aged 8 weeks and weighing 20-22 g were provided by the Experimental Animal Center of Zunyi Medical University. All animals were housed in a barrier system at the Experimental Animal Center with a temperature of 20-26°C, a humidity of 50±5%, and a 12-hour day-night cycle. All animal experiments were conducted in accordance with the Guide for the Care and Use of Laboratory Animals, and all animal experiments were approved by the Experimental Animal Welfare Ethics Committee of Zunyi Medical University.

[0054] 1.2 Experimental methods

[0055] 1.2.1 Preparation and quality control of cinnabar root water extract

[0056] Take 200g of cinnabar root and add water (10 times the amount) for heating reflux extraction twice, each time for 2 hours, combine the filtrate, filter and concentrate to 1g / mL (based on crude drug amount), and store at -20℃; prepare the reference solution and the test solution according to the 2020 edition of the Chinese Pharmacopoeia, accurately draw 5μL of the reference solution and the test solution, inject them into the liquid chromatograph and measure, and finally obtain the HPLC chart ( Figure 1 ). As can be seen from the figure, both can detect bergenin at a wavelength of 275nm, which meets the pharmacopoeia standards.

[0057] 1.2.2 Construction of experimental animal models

[0058] Periodontitis: 50 8-week-old C57BL / 6JSPF grade male mice were fed adaptively for 1 week, and 40 mice were randomly selected for anesthesia, fixed in the supine position, and the tongue was pulled out for fixation. A 1mL injection needle was used to separate the mesiodistal interproximal space of the maxillary second molars on both sides, and 5-0 silk thread was passed through the mesiodistal interproximal space on both sides in turn, and circled the tooth neck, and finally tied on the buccal side. The gingival state, tooth looseness and silk thread shedding were observed every other day, and if the silk thread fell off, it was ligated in time.

[0059] Ulcerative colitis: 20 mice were given 3% dextran sulfate sodium salt (DSS) solution to drink freely for 5 days, and fresh DSS solution was replaced every other day to induce mouse ulcerative colitis model (see Figure 2 ).

[0060] 1.2.3 Determination of the dosage of cinnabar root and grouping of experimental animal models

[0061] The experimental dose of cinnabar root was determined with reference to the 2020 edition of the Chinese Pharmacopoeia. The pharmacopoeia stipulates that the daily dosage for humans is 3 to 9 g, and the dosage per unit body weight is converted into a mouse dosage of 0.39 to 1.17 g / kg. In order to evaluate the anti-inflammatory effect of cinnabar root, a periodontitis model was constructed for 12 mice, which were randomly divided into 3 groups of 4 mice each. 2, 4, and 8 times the dose (0.78, 1.56, and 3.12 g / kg) were selected for administration, and the optimal dose was finally screened out by ELISA experiment for subsequent experiments.

[0062] Fifty male mice were randomly divided into five groups, with 10 mice in each group, namely the normal control NC group, the periodontitis P group, the periodontitis and colitis PU group, the cinnabar root treatment periodontitis PD group, and the cinnabar root treatment periodontitis and colitis PUD group. The experimental animal model was established according to the method in 1.3.6. Three days after the periodontitis model was established, the PD group and the PUD group began to receive the cinnabar root by gavage, and the other groups were gavaged with distilled water, as shown in Figure 1. Figure 3 The growth status and weight changes of mice in each group were observed every day, and the fecal bleeding of mice in the PU group and PUD group was evaluated and scored according to the disease activity index (DAI) scoring standard in Table 1.

[0063] Table 1 DAI scoring criteria

[0064]

[0065] 1.2.4 Sample collection and processing

[0066] Fecal sample collection was controlled at the same time of the day. Combined with the natural defecation and stress defecation of mice, disposable sterile instruments were used to collect the feces of each group of mice, placed in sterile and enzyme-free cryopreservation tubes, and after quick freezing with liquid nitrogen, they were immediately stored at -80℃ for later use.

[0067] After the mice were killed by cervical dislocation, various tissue samples were collected. The specific collection results are shown in Table 2.

[0068] Table 2 Sample collection and processing

[0069]

[0070] 1.2.5 Metabolomics research

[0071] In order to clarify the metabolic regulatory mechanism of cinnabar root on periodontitis and colitis, the present invention collected the feces of mice on the 14th day (after the periodontitis modeling was completed) and the 21st day (after the periodontitis and colitis modeling were completed) to detect the changes in metabolites in the mouse feces, and comprehensively investigated the effects of cinnabar root on the metabolism of periodontitis and colitis.

[0072] (1) Sample pretreatment

[0073] Accurately weigh an appropriate amount of sample into a 2 mL sterile centrifuge tube, add 600 μL of methanol containing 2-chloro-L-phenylalanine (4 ppm), vortex for 30 seconds and mix thoroughly; add steel beads and put into a high-throughput tissue grinder for grinding at 50 Hz for 120 seconds; ultrasonicate at room temperature for 10 minutes; centrifuge at 12,000 rpm and 4°C for 10 minutes, take the supernatant and filter it using a 0.22 μm membrane, and add the filtrate to the detection bottle for LC-MS detection.

[0074] (2) Chromatographic conditions

[0075] use HSST3 (2.1×100 mm, 1.8 μm) column, flow rate of 0.3 mL / min, column temperature of 40°C, injection volume of 2 μL. Positive ion mode, mobile phase B2 is 0.1% formic acid acetonitrile, mobile phase A2 is 0.1% formic acid water, gradient elution program is: 0-1 min, 8% B2; 1-8 min, 8%-98% B2; 8-10 min, 98% B2; 10-10.1 min, 98%-8% B2; 10.1-12 min, 8% B2. In negative ion mode, the mobile phase was acetonitrile (B3) and 5 mM ammonium formate water (A3), and the gradient elution program was: 0-1 min, 8% B3; 1-8 min, 8%-98% B3; 8-10 min, 98% B3; 10-10.1 min, 98%-8% B3; 10.1-12 min, 8% B3.

[0076] (3) Mass spectrometry conditions

[0077] The positive ion spray voltage was 3.50 kV, the negative ion spray voltage was -2.50 kV, the sheath gas was 40 arb, and the auxiliary gas was 10 arb. The capillary temperature was 325 °C, and the primary full scan was performed at a resolution of 70,000. The primary ion scan range was m / z 100-1000, and HCD was used for secondary fragmentation. The collision energy was 30 eV, the secondary resolution was 17,500, and the first 10 ions of the acquisition signal were fragmented. At the same time, dynamic exclusion was used to remove unnecessary MS / MS information.

[0078] (4) Metabolomics data processing

[0079] Quality control (QC) sample analysis is a necessary step to examine the stability and accuracy of the instrument metabolomics results. Support vector regression correction based on QC samples is used to eliminate systematic errors, and substances with RSD>30% in QC samples are filtered out during the quality control and quality assurance process for subsequent data analysis. Public databases such as HMDB, Massbank, LipidMaps, mzCloud, KEGG and other spectral databases are used for substance identification, and the parameters are set to ppm<30ppm to obtain qualitative results of metabolites.

[0080] (5) Metabolomics data analysis

[0081] The R software package Ropls was used to perform PLS-DA and OPLS-DA dimensionality reduction analysis on the sample data. The t-test method was used to test the overfitting of the model. The P value was calculated based on the statistical test, the variable projection importance (VIP) was calculated by the OPLS-DA dimensionality reduction method, and the fold change was used to calculate the difference between groups to measure the influence and explanatory power of each metabolite content on the classification and discrimination of samples, and to assist in the screening of marker metabolites. Combined with P<0.05 and VIP>1, differential metabolites were screened, and the MetaboAnalyst software package was used to perform functional pathway enrichment and topological analysis on the screened differential metabolites.

[0082] 1.2.6 Transcriptomics studies

[0083] (1) Sample pretreatment

[0084] The RNA of mouse colon tissue was extracted using the Trizol kit. The extracted RNA samples were then subjected to strict quality control, which mainly included the following two aspects: first, agarose gel electrophoresis was used to analyze the integrity of the RNA sample and whether there was DNA contamination; second, the Nanodrop micro-spectrophotometer was used to perform preliminary quantification of RNA to detect concentration and purity.

[0085] (2) Transcriptome sequencing

[0086] Transcriptome sequencing is the sequencing analysis of all mRNAs. The specific operation process is as follows Figure 4 shown.

[0087] (3) Transcriptome data analysis

[0088] The original image files were converted into sequencing reads by CASAVA base recognition. In order to ensure the quality and reliability of data analysis, the original data needed to be filtered, that is, reads with adapters were removed; reads with N (N means that the base information cannot be determined) ratio greater than 0.002 were removed; when the number of low-quality bases contained in the single-end read exceeded 50% of the length ratio of the read, the paired reads were removed. The sequencing error rate and GC content distribution were checked, and finally the clean reads used for subsequent analysis were obtained. The RNA-Seq sequencing data were compared and analyzed using Hisat2 software.

[0089] HTSeq software was used to quantitatively analyze the FPKM (Fragments Per Kilobase of transcriptsequence per Millions base pairs sequenced) value. FPKM is the number of fragments per kilobase length from a gene / transcript per million fragments, which takes into account the effects of sequencing depth and gene length on fragment counts. The edgeR software was used to perform expression difference significance analysis, and P<0.05 was used as the difference significance standard. ClusterProfiler software was used to select the widely used annotation gene databases GO, KEGG and Reactome; GSEA enrichment analysis was used to identify differentially regulated pathways between the PU group and the PUD group.

[0090] 1.2.7 Real-time fluorescence quantitative reverse transcription PCR

[0091] (1) Extraction of RNA from colon tissue

[0092] ① Weigh 100 mg of fresh colon tissue, wash it with PBS, place it in a 1.5 mL sterile enzyme-free EP tube, add 1 mL of Trizol reagent and 3 sterile enzyme-free zirconium oxide grinding beads, grind it thoroughly using a high-throughput tissue grinder, and place it on ice for 5 minutes to allow it to be fully lysed.

[0093] ② Add 200 μL of chloroform and shake thoroughly to mix. After standing for 5 minutes, centrifuge at 12,000 rpm for 10 minutes at 4°C, aspirate the top aqueous phase, and transfer the supernatant to a new sterile, enzyme-free EP tube.

[0094] ③ Add 0.5 to 1 times the amount of isopropanol as the supernatant and mix well. Place on ice for 20 minutes, centrifuge at 12,000 rpm for 15 minutes at 4°C, and discard the waste liquid.

[0095] ④ Add 1 ml of 75% ethanol to wash the RNA precipitate. After gentle shaking, centrifuge at 12,000 rpm for 10 min at 4°C and discard the waste liquid (repeat once).

[0096] ⑤After vacuum drying the RNA precipitate for 5 minutes, add 50 μL DEPC water to dissolve it.

[0097] (2) RNA reverse transcription

[0098] Table 3 Reverse transcription reaction system

[0099]

[0100] Table 4 Reaction conditions

[0101]

[0102] (2) cDNA amplification

[0103] Table 5 Reverse transcription reaction system

[0104]

[0105] Table 6 Primer sequences

[0106]

[0107] 1.3 Statistical methods

[0108] SPSS 29.0 (IBM Corp., Armonk, NY, USA, Version 29.0) was used for data analysis. The measured data were expressed as mean ± standard deviation. When normality and homogeneity of variance were met, t-test was used for pairwise comparison. One-way ANOVA was used for analysis of variance between different groups. P < 0.05 was considered statistically significant.

[0109] 2 Results

[0110] 2.1 Therapeutic effect of cinnabar root on model mice

[0111] 2.1.1 Determination of the dosage of cinnabar root

[0112] The expression level of inflammatory factor IL-1β in serum was detected by ELISA experiment, and it was found that the best effect of inhibiting inflammatory factors was achieved when the dosage was 1.56 g / kg (Table 7). Therefore, 1.56 g / kg was selected for subsequent experiments.

[0113] Table 7 Mouse serum IL-1B concentration

[0114]

[0115]

[0116] Note: Data is based on Indicates, one-way ANOVA (n=4); ***P<0.001, **P<0.01, *P<0.05

[0117] 2.1.2 Growth status and weight changes of mice in each group

[0118] Through observation, it was found that all groups of mice could tolerate the establishment of periodontitis and colitis models and the intervention of cinnabar root. The hair of mice in the NC, P and PD groups was smooth and dense, they were energetic and vigorous, and their diet and sleep were normal. Compared with the NC group, the mice in each group induced by DSS solution showed poor vitality, loss of appetite, loose and rough hair, and slow behavior. Three days after modeling, they began to show intestinal symptoms such as fecal occult blood, diarrhea, and weight loss to varying degrees. After the intervention of cinnabar root, compared with the PU group model mice, the PUD group mice improved the symptoms of DSS-induced colitis in mice, reduced the DAI score, and the mental state of the mice was also restored ( Figure 5 ).

[0119] 2.1.3 Periodontal tissue pathological analysis and alveolar bone height changes

[0120] HE staining results showed that compared with the NC group, the integrity and continuity of the gingival epithelium in the model group P group and PU group were irregularly destroyed and extended to the root direction, edema degeneration, subepithelial connective tissue was densely infiltrated by inflammatory cells, alveolar bone and intrinsic alveolar ridge top were destroyed and absorbed to varying degrees, and obvious gingival recession was observed. After intervention with cinnabar root, compared with the model group P group and PU group, the PD group and PUD group had a protective effect on periodontal tissue, reduced the infiltration of inflammatory cells, and restored the height of part of the alveolar ridge ( Figure 6 ).

[0121] After Micro-CT scanning of mouse maxillary bone samples for reconstruction, the degree of alveolar bone resorption in each group was compared. The 3D and 2D reconstruction images showed that compared with the NC group, the maxillary second molars in the P and PU groups had obvious alveolar bone resorption, and the CEJ-ABC distance increased, indicating that the experimental periodontitis was successfully constructed. Compared with the P and PU groups, the alveolar bone resorption in the PD and PUD groups was reduced, and the CEJ-ABC distance was reduced, which was statistically significant (P<0.05). Figure 7 ).

[0122] 2.1.4 Colonic histopathological analysis and changes in tight junction proteins

[0123] The results of HE staining of colon tissue showed that mucosal damage was observed in the colon tissue of the PU group after DSS induction, accompanied by a large number of inflammatory cell infiltrations, reduced or even disappeared mucus in goblet cells, abnormal crypt structure, and irregular surface. After treatment with cinnabar root, the PUD group reversed the above symptoms and alleviated colon inflammation ( Figure 8 ). The expression levels of tight junction proteins ZO-1 and Occludin in each group were detected by immunofluorescence staining. It was found that compared with the model group, the drug group after cinnabar root intervention increased the expression levels of tight junction proteins ZO-1 and Occludin, suggesting that cinnabar root can repair the damaged intestinal barrier ( Fig. 9 ).

[0124] 2.1.5 Histopathological analysis of heart, liver and kidney

[0125] The myocardial cells, hepatocytes and glomeruli of mice in the NC group were intact and clear, with orderly arrangement of cells and uniform staining. The myocardial cells, hepatocytes and glomeruli of mice in the model group, P group and PU group were disorderly arranged, myocardial fibers were broken, and the hepatocytes and glomeruli were abnormally structured, accompanied by varying degrees of inflammatory infiltration. Compared with the model group, the PD group and PUD group after drug treatment had significant improvements in the morphology and structure of myocardial cells, hepatocytes and glomeruli, with orderly arrangement of cells and a decrease in inflammatory cells ( Fig.10 ).

[0126] 2.1.6 Effects of cinnabar root on inflammatory factors in model mice

[0127] To investigate the potential effects of cinnabarinum root on the inflammatory response of periodontitis-associated colitis, the expression levels of inflammatory factors in serum and colon tissue were evaluated ( Fig.11 Compared with the NC group, the P and PU groups significantly increased the expression levels of pro-inflammatory cytokines TNF-α, IL-1β, and IL-6 in serum and colon, and decreased the expression level of anti-inflammatory factor IL-10; however, after treatment with cinnabar root, compared with the P and PU groups, the expression levels of TNF-α, IL-1β, and IL-6 in serum and colon of the PD and PUD groups were significantly decreased, and the expression level of IL-10 was significantly increased, and the differences were statistically significant (***P<0.001, **P<0.01, *P<0.05).

[0128] 2.2 Effects of cinnabar root on fecal metabolomics of model mice

[0129] 2.2.1 Metabolic profiling analysis

[0130] The present invention collects the feces of mice on the 14th and 21st day to detect the metabolic changes in the feces of mice. Fig.12 ) and OPLS-DA( Fig.13) method was used to analyze the metabolic profiles of feces in positive and negative ion modes. The results showed that on the 14th day, the fecal samples of the NC1 group and the P1 group, and the P1 group and the PD1 group were clustered within the 95% confidence interval and the separation degree of each group was significant, indicating that there were significant differences in fecal metabolites between the NC1 group and the P1 group, and the P1 group and the PD1 group. After induction with DSS solution, the fecal samples of the P2 group and the PU2 group were analyzed on the 21st day and found that the two groups were each on one side, indicating that the type or level of metabolites was changed after the successful establishment of the periodontitis with colitis model, and after the intervention of cinnabar root administration, the clustering differences between the PUD2 group and the PU2 group were obvious, suggesting that cinnabar root may alleviate the progression of periodontitis with colitis by regulating the changes in metabolic profiles.

[0131] 2.2.2 Screening and cluster analysis of differential metabolites

[0132] The contribution of each group in the OPLS-DA analysis was screened according to the VIP value, and the metabolite markers associated with the treatment of periodontitis and colitis with cinnabar root were screened. When the VIP value was greater than 1.0 and P<0.05, it indicated that the metabolite had a large contribution and a high influence, and could be selected as a primary differential metabolite marker; a total of 7,947 differential metabolites were initially screened, of which 1,167 candidate differential metabolites were found between the NC1 group and the P1 group, 76 candidate differential metabolites were found between the P1 group and the PD1 group, 302 candidate differential metabolites were found between the P2 group and the PU2 group, and 27 candidate differential metabolites were found between the PU group and the PUD group. Fig.14 shown.

[0133] The identified candidate differential metabolites were searched and compared with spectral databases such as HMDB, Massbank, LipidMaps, mzCloud, and KEGG, and a total of 673 metabolites were finally screened out, including down-regulated tryptophan and ethyl guanidinoacetate in the NC1 and P1 groups, and up-regulated succinic acid, β-guanidinopropionic acid, 4-hydroxystyrene, and 2,4-dihydroxyquinoline; down-regulated 4-hydroxystyrene, 2,4-dihydroxyquinoline, and acetic acid in the P1 and PD1 groups. D-glucono-1,4-lactone, up-regulated 13S-hydroxyoctadecadienoic acid, dicyclomine, p-aminobenzoic acid, dibutyl phthalate, caproic acid, etc.; P2 group and PU2 group included up-regulated benzaldehyde, 5-aminovaleric acid, 4-hydroxystyrene, down-regulated L-proline, adenine, etc.; PU group and PUD group included down-regulated 2,4-dihydroxyquinoline, D-glucono-1,4-lactone, ribavirin, and up-regulated 13S-hydroxyoctadecadienoic acid, dicyclomine, misoprostol, etc. It is suggested that the main differential metabolites of cinnabar root in the treatment of periodontitis with colitis include 2,4-dihydroxyquinoline, D-glucono-1,4-lactone, 13S-hydroxyoctadecadienoic acid, dicyclomine, etc. In order to more intuitively illustrate the differences between the groups, the above metabolites were drawn into a heat map, as shown in Fig.15 , Fig.16 shown.

[0134] The metabolism of substances in the body is mutually regulated to maintain stability. Different metabolites have cooperative or mutually exclusive relationships. The correlation between each metabolite is analyzed by calculating the Pearson correlation coefficient between the two differential metabolites. The metabolites with significant correlation are statistically tested and analyzed. Metabolites with significant P<0.05 are selected for mapping ( Fig.17 ).

[0135] 2.2.3 Enrichment analysis of metabolic pathways

[0136] First, the significantly different metabolites identified in each group of feces were integrated, and then the sorted differential metabolites were imported into the Metaboanalyst database. The enrichment method was based on the hypergeometric distribution test, and the topological analysis used the degree centrality method to screen the metabolic pathways involved in the above differential metabolites ( Fig.18 ). Compared with NC1, the early periodontitis P1 group had alterations in metabolic pathways such as central carbon metabolism, arginine biosynthesis, arginine and proline metabolism, prolactin signaling pathway, D-arginine and D-ornithine metabolism, prostate cancer, phenylalanine metabolism, alanine, aspartate, and glutamate metabolism ( Fig.18 A); Under the intervention of cinnabar root, compared with the P1 group, the PD1 group showed changes in metabolic pathways such as PPAR signaling pathway, linoleic acid metabolism, galactose metabolism, lysine degradation, retrograde endocannabinoid signaling, biosynthesis of phenylalanine, tyrosine and tryptophan, and biosynthesis of valine, leucine and isoleucine ( Fig.18 B). After induction with DSS solution, the metabolic pathways of the P2 group were changed in ABC transporters, prostate cancer, vitamin digestion and absorption, pathways in cancer, linoleic acid metabolism, phenylalanine, tyrosine and tryptophan biosynthesis, etc. compared with the PU2 group ( Fig.18 C); after intervention with cinnabar root, compared with the PU2 group, the PUD2 group showed changes in linoleic acid metabolism, PPAR signaling pathway, unsaturated fatty acid biosynthesis pathway, bile secretion, steroid hormone biosynthesis, thiamine metabolism, phenylalanine metabolism and ABC transporter pathway ( Fig.18 D). These results suggest that cinnabar root may effectively regulate metabolic disorders in periodontitis and colitis mainly through PPAR signaling pathway, phenylalanine and linoleic acid metabolism.

[0137] 2.3 Effects of cinnabar root on colon transcriptomics in model mice

[0138] 2.3.1 Sequencing data quality assessment

[0139] After filtering the raw data, checking the sequencing error rate and GC content distribution, clean reads were obtained for subsequent analysis, and the data are summarized in Table 8. The results showed that the alignment rate of sequencing reads (Total mapped) of each group of samples was higher than 95%, and the number of reads with multiple alignment positions on the reference sequence (multiple mapped reads) accounted for less than 12%, indicating that the sample results were credible and suitable for subsequent analysis.

[0140] Table 8 Data quality summary

[0141]

[0142] 2.3.2 Principal Component Analysis

[0143] Principal Component Analysis (PCA) is often used to evaluate inter-group differences and intra-group sample duplication. PCA uses linear algebra calculation methods to reduce the dimension and extract principal components of tens of thousands of genetic variables. The results of PCA analysis showed that the samples between the NC group and the PU group were dispersed, each in one category, indicating that the transcriptional profiles between the periodontitis with colitis model after successful establishment were significantly different from those of the normal control group; after intervention with cinnabar root, the samples between the PU group and the PUD group were significantly separated, suggesting that cinnabar root may prevent and treat periodontitis with colitis by regulating gene transcription ( Fig.19 ).

[0144] 2.3.3 Screening and cluster analysis of differentially expressed genes

[0145] The edgeR software was used to analyze the significance of gene expression differences, and P<0.05 was used as the standard for difference significance. The volcano plot was used to intuitively display the overall distribution of genes with significant expression differences. Compared with the normal group NC, there were 2,155 significantly differentially expressed genes in the periodontitis with colitis group PU, of which 1,128 genes were significantly upregulated and 1,027 genes were significantly downregulated, mainly including Il17a, Cdc45, H19, Il17f, Il6, Cd52, etc. Compared with the model group PU, there were 230 significantly differentially expressed genes in the PUD group after cinnabar root drug intervention, of which 76 genes were significantly upregulated and 154 were significantly downregulated. The key differentially expressed genes included Socs3, Gsta3, Ptk6, Col1a1, Tat, Fkbp11, Il6, etc. ( Fig. 20 ). The mainstream hierarchical clustering method was used to convert the log10(FPKM+1) value (scale number) and perform clustering to obtain the differential gene hierarchical clustering heat map ( Fig.21 ).

[0146] 2.3.4 GO enrichment analysis of differentially expressed genes

[0147] GO enrichment analysis of the NC and PU groups yielded a total of 855 entries (P<0.05), including 551 biological process (BP) entries, 136 cell composition entries (CC), and 168 molecular function entries (MF). The top ten entries were selected for mapping, including innate immune response, immunoglobulin production, DNA replication, response to bacteria, extracellular region, plasma membrane outer side, plasma membrane, circulating immunoglobulin complex, immunoglobulin receptor binding, cytokine activity, antigen binding, and same protein binding ( Fig. 22 A); GO enrichment analysis of the PU group and the PUD group obtained a total of 332 entries (P<0.05), including 235 biological process (BP) entries, 24 cell composition entries (CC), and 73 molecular function entries (MF), mainly including acute phase response, response to bacteria, innate immune response, B cell receptor signaling pathway, inflammatory response, extracellular region, plasma membrane outer side, plasma membrane, immunoglobulin complex, cytokine activity, oxidoreductase activity and glutathione transferase activity ( Fig. 22 B).

[0148] 2.3.5 KEGG pathway and GSEA enrichment analysis and verification of differentially expressed genes

[0149] KEGG pathway enrichment analysis between the NC group and the PU group found that a total of 1,427 differentially expressed genes were involved in regulation, and 95 significantly related pathways were enriched (P<0.05), mainly including CD molecules, cytokine-cytokine receptor interaction, DNA replication, NOD-like receptor signaling pathway, cell cycle and IL-17 signaling pathway ( Fig.23 A). KEGG pathway enrichment analysis of the PU group and the PUD group yielded 52 significantly correlated pathways (P<0.05), including cytokine-cytokine receptor interaction, drug metabolism, hematopoietic cell lineage, IL-17 signaling pathway, JAK-STAT signaling pathway, cytochrome P450, TNF signaling pathway, platinum resistance, and linoleic acid metabolism ( Fig.23 B). Integrative analysis revealed that the IL-17 signaling pathway and the JAK-STAT signaling pathway may be the key pathways for the intervention of cinnabar root with periodontitis and colitis, and the differentially regulated pathways between the groups were also verified by GSEA enrichment analysis. The IL-17 signaling pathway and the JAK-STAT signaling pathway were downregulated between the PU and PUD groups (P<0.01)( Fig.24); In addition, the expression changes of IL-17A, IL-17F, JAK2 and STAT3, the key genes of the pathway, in the PU and PUD groups were detected by real-time fluorescence quantitative PCR technology. It was found that after intervention with cinnabar root, the expression levels of IL-17A, IL-17F, JAK2 and STAT3 were inhibited ( Fig.25 ).

[0150] 3 Discussions

[0151] 3.1 Transcriptomics study of cinnabar root and its effects on periodontitis-associated colitis

[0152] Cinnabar root is a traditional Chinese medicine of the Miao ethnic group. Its root has high medicinal value and has been used clinically to treat respiratory tract infections, colitis, acute lung injury, bronchitis, bladder cancer, cervical cancer and many other diseases. Kaihoujian spray is an exclusive Miao medicine produced by Guizhou Sanli Pharmaceutical Co., Ltd. Cinnabar root plays a key role as its formula medicinal material; the main effective components of cinnabar root include bergenin, triterpenoid saponins and flavonoids, among which bergenin, as an indicator component of cinnabar root medicinal materials in the "Chinese Pharmacopoeia", also has anti-inflammatory, anti-tumor and antioxidant effects. In recent years, with the increasing use of cinnabar root in the private sector and pharmaceutical companies, the market demand has been greater, resulting in the emergence of mixed counterfeit products with the same name as cinnabar root on the market, resulting in obvious differences in the quality of cinnabar root medicinal materials, which seriously affects the safety of clinical precision medication of cinnabar root and the effectiveness of quality evaluation of traditional Chinese medicine. At present, due to the similarity of the appearance of the cinnabar root and its closely related species, and the susceptibility to subjective factors, species identification cannot be carried out well. However, DNA barcoding technology can make up for some of the shortcomings of traditional identification methods at the molecular level. This technology can easily and quickly identify a large number of samples at one time, and even if the samples are damaged, it will not affect the identification results. It has the characteristics of high throughput and high accuracy, and has been widely used in the identification research of various species.

[0153] There is a certain correlation between periodontitis and colitis. The bacterial pathogens produced in periodontitis can aggravate the occurrence and development of colitis. Similarly, the occurrence of colitis can also affect the health of periodontal tissues. Both periodontitis and colitis are chronic inflammatory diseases. They share some common pathophysiological mechanisms, which are mainly related to the disorder of host-microbiota interaction and immune regulation disorder. The oral-gut axis mediated by the host and microbiota is currently a hot topic of concern. The proinflammatory immune cells produced by periodontitis can migrate to the gastrointestinal tract through the oral-gut axis, thereby affecting intestinal immunity and inflammatory response. The mechanisms regulating the connection between oral and systemic health include susceptibility and inducing factors, such as genetic factors, environmental factors, and drugs. The occurrence and development of periodontitis and colitis may be affected by genetic and environmental factors. Studies have found that periodontitis activates colitis inflammasomes, changes the composition and metabolite profile of the intestinal microbiota of colitis mice, and can even promote the occurrence of colorectal cancer. The meta-analysis also found that the diagnosis of inflammatory bowel disease was associated with the risk of periodontitis. Compared with the healthy control group, IBD patients had significantly more patients with moderate to severe periodontitis (85.6% vs. 65.6%, p<0.001) and severe periodontitis (36.7% vs. 25.6%, p<0.001). After subgroup analysis, patients with ulcerative colitis had a higher risk of periodontitis than patients with Crohn's disease.

[0154] GO enrichment analysis of differentially expressed genes based on transcriptomics (P<0.05) showed that the effects of cinnabar root on periodontitis with colitis were mainly in the acute phase response, response to bacteria, innate immune response, B cell receptor signaling pathway, inflammatory response, extracellular region, plasma membrane outer side, plasma membrane, immunoglobulin complex, cytokine activity, oxidoreductase activity and glutathione transferase activity. KEGG pathway enrichment analysis showed that cinnabar root treatment of periodontitis with colitis enriched 52 significantly related pathways (P<0.05), mainly including cytokine-cytokine receptor interaction, drug metabolism, hematopoietic cell lineage, IL-17 signaling pathway, JAK-STAT signaling pathway, cytochrome P450, TNF signaling pathway, platinum resistance and linoleic acid metabolism. GSEA enrichment analysis of differentially regulated pathways between the groups showed that the IL-17 signaling pathway and JAK-STAT signaling pathway in periodontitis with colitis were reduced after drug intervention with cinnabar root (P<0.01). Based on the above transcriptomic results, integrated analysis suggested that cinnabar root may intervene in the treatment of periodontitis with colitis through the IL-17 signaling pathway and the JAK-STAT signaling pathway.

[0155] In experimental periodontitis, the gene expression levels of IL17F, IL17A, and STAT3 increased, promoting alveolar bone loss. Similarly, osteoclast-derived exosomes miR-5134-5p interfered with alveolar bone homeostasis by targeting the JAK2 / STAT3 axis; IL-17 can inhibit iron concentration through the interaction between NRF2 and p-STAT3, alleviating ferroptosis-induced alveolar bone loss. In addition, STAT3 is a key gene linking inflammation and cancer. Whole-genome analysis of inflammatory bowel disease found that STAT3 is also one of the susceptibility genes for inflammatory bowel disease; the inflammatory factor IL-6 can activate STAT3 in colon epithelial cells (IECs), thereby promoting the activation of the ceramide and sphingosine phosphate 1 pathways, changing the recruitment of immune cells CD8+T and Treg, promoting cell proliferation, and ultimately leading to the occurrence of colon cancer. In view of this, the present invention detected the expression changes of key genes IL-17A, IL-17F, JAK2 and STAT3 in the IL-17 signaling pathway and JAK-STAT signaling pathway in the PU and PUD groups by real-time fluorescence quantitative PCR technology, and found that the expression levels of IL-17A, IL-17F, JAK2 and STAT3 were reduced after treatment with cinnabar root.

[0156] 3.2 Metabolomics study of cinnabar root on periodontitis and colitis

[0157] Non-targeted metabolomics can reveal key metabolites and their association with diseases to discover new biomarkers, metabolic pathways and drug targets. The present invention explores the metabolic change pathways of cinnabar root on periodontitis with colitis by collecting the feces of mice on the 14th and 21st days. A systematic analysis was conducted from the early periodontitis and blank control groups, to periodontitis and periodontitis with colitis, and then to the experimental animal model of cinnabar root drug intervention. The results showed that cinnabar root mainly effectively regulated periodontitis with colitis through differential metabolites such as 2,4-dihydroxyquinoline, D-glucono-1,4-lactone, 13S-hydroxyoctadecadienoic acid, and dicyclomine. Alkylquinolones have been proven to be antibacterial drugs. At present, 2,4-dihydroxyquinoline has been synthesized as a new drug, and it has been found that it significantly affects the antibacterial activity. Linoleic acid metabolites produced by intestinal flora may affect host lipid metabolism by regulating peroxisomal β-oxidation activity. Among them, 13S-hydroxyoctadecadienoic acid, as a derivative of linoleic acid, plays a role in regulating inflammation and tumor development in different cell systems (such as endothelial cells); in the proximal colon of UC, the abundance of 13S-hydroxyoctadecadienoic acid is reduced, and its abundance is adjusted after drug treatment, which is consistent with the results of the present invention. Cinnabar root increases the abundance of 13S-hydroxyoctadecadienoic acid in periodontitis with colitis. Dicyclomine is an anticholinergic that has been used to treat gastrointestinal diseases such as acid gastrointestinal disease and irritable bowel syndrome.

[0158] The results of KEGG enrichment of differential metabolites showed that cinnabar root effectively regulated metabolic disorders in periodontitis and colitis mainly through PPAR signaling pathway, phenylalanine metabolism, linoleic acid metabolism, unsaturated fatty acid biosynthesis pathway, bile secretion, steroid hormone biosynthesis and other pathways. Peroxisome proliferators-activated receptors (PPAR) are divided into three subtypes: PPAR-α (NR1C1), PPAR-δ / β (NR1C2) and PPAR-γ (NR1C3), which can regulate a variety of biological processes, including inflammation, lipid and glucose metabolism, and overall energy balance. PPAR is also related to the normal homeostasis of intestinal tissue. In UC, the mRNA expression of PPAR-γ is reduced, indicating that PPAR is negatively correlated with the progression of UC; in addition, PPAR can improve metabolic disorders when combined with agonist fibrates. Rosiglitazone inhibits inflammatory responses by regulating PPAR-γ and downregulating the TLR / NF-κB signaling pathway, thereby alleviating or preventing Porphyromonas gingivalis-accelerated atherosclerosis. The results of non-targeted metabolomics in the serum of mice fed an isoflavone diet showed that the synthesis of anti-inflammatory unsaturated fatty acids increased, while the pro-inflammatory phenylalanine metabolism and arachidonic acid derivatives decreased. Therefore, it is speculated that the treatment of periodontitis with colitis by cinnabar root may be achieved by inhibiting inflammatory factors through the PPAR signaling pathway, promoting the biosynthesis of unsaturated fatty acids, inhibiting phenylalanine metabolism, and increasing the abundance of 2,4-dihydroxyquinoline and linoleic acid metabolic derivative 13S-hydroxyoctadecadienoic acid.

[0159] 4. Conclusion

[0160] (1) Cinnabar root can effectively treat periodontitis with colitis by inhibiting inflammatory factors and repairing damaged barrier ZO-1 and Occludin.

[0161] (2) Based on the integrated analysis of metabolomics and transcriptomics, it was shown that cinnabar root may effectively treat periodontitis with colitis by inhibiting the IL-17 signaling pathway, the JAK-STAT signaling pathway, affecting the PPAR signaling pathway, linoleic acid metabolism, the biosynthesis of unsaturated fatty acids, phenylalanine metabolism, and regulating immune response.

[0162] (3) By tapping into the resource advantages of the local "authentic medicinal materials" industry brand, it will be beneficial to the scientific proof of cinnabar root, promote the promotion and application of cinnabar root authentic medicinal materials in Guizhou Province, and then carry forward and inherit the characteristic ethnic medicine of Guizhou Province.

Claims

1. Use of cinnabar root or its extract in preparing medicine for treating periodontitis.

2. Use of cinnabar root or its extract in the preparation of a medicine for treating ulcerative colitis.

3. Use of cinnabar root or its extract in the preparation of medicines for treating periodontitis with colitis.

4. The use according to any one of claims 1 to 3, characterized in that: The cinnabari root extract is a cinnabari root water extract.

5. The use according to claim 4, characterized in that: The preparation method of the cinnabar root water extract is as follows: adding water to the cinnabar root, heating and refluxing for extraction, combining the filtrate, filtering and concentrating to 1 g / mL (based on crude drug amount), and obtaining the cinnabar root water extract.

6. The use according to any one of claims 3 to 5, characterized in that: The drug has the functions of down-regulating inflammatory factors, repairing damaged barriers ZO-1 and Occludin, inhibiting IL-17 signaling pathway, JAK-STAT signaling pathway, affecting PPAR signaling pathway, linoleic acid metabolism, and regulating immune response.

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