Application of SLC9A1 protein and S100A2 protein as action targets in preparation of medicine for treating primary hypertension

Abstract

The invention relates to the field of biological pharmacy, in particular to application of SLC9A1 protein and S100A2 protein as action targets in preparation of drugs for treating primary hypertension. The invention provides an application of an SLC9A1 protein and an S100A2 protein as action targets in preparation of a medicine for treating primary hypertension. In the specific embodiment of the invention, after the MMBIT treatment is carried out on the EH patient, the result shows that the SLC9A1 protein and the S100A2 protein in the blood of the EH patient are obviously improved after the MMBIT treatment, and finally the aim of improving the systolic pressure and the diastolic pressure of the EH patient is achieved. Therefore, the SLC9A1 protein and the S100A2 protein can serve as action targets to be used for preparing the medicine for treating the primary hypertension, and a new treatment way is provided for treatment of the EH.

Description

Technical Field

[0001] The present invention relates to the field of biopharmaceuticals, and in particular to the use of SLC9A1 protein and S100A2 protein as action targets in the preparation of drugs for treating essential hypertension. Background Art

[0002] Hypertension (essential hypertension, EH) is defined as a systolic blood pressure higher than 140 mmHg and a diastolic blood pressure higher than 90 mmHg. EH is the most important modifiable risk factor for cardiovascular and kidney diseases, and the main risk factor for death. It poses a major public health challenge to both economically developing and developed countries. Compared with patients with normal blood pressure, EH patients have a higher risk of cardiovascular and cerebrovascular diseases, and the prevalence of EH increases significantly with age, and the trend of younger patients is becoming increasingly significant. The blood vessel itself is a complex organ that can autonomously produce local active substances. Changing the physical properties of the blood vessels can easily cause significant changes in individual blood pressure.

[0003] The occurrence and development of any disease is not only manifested in the body, but also in the mind. In 1976, Esler M believed that EH may be caused by a psychosomatic mechanism. Some national health departments stated that depression can be considered a cardiac risk factor. EH patients often show negative psychological and behavioral characteristics such as emotional tension, depression, anger, and anxiety. Whether it is social and psychological stress factors, emotional factors or behavioral factors, they can increase blood pressure and induce EH.

[0004] At present, the treatment of EH mainly relies on taking antihypertensive drugs. Different antihypertensive drugs are taken according to the pathogenesis of EH. Despite the wide variety of antihypertensive drugs, only one in six EH patients can effectively control their blood pressure, but there will be certain side effects (symptoms such as headache, edema and bradycardia). Therefore, it is necessary to find a method that has no side effects and is effective in treating EH. Summary of the invention

[0005] The purpose of the present invention is to provide the use of SLC9A1 protein and S100A2 protein as targets in the preparation of drugs for treating essential hypertension, so as to solve the problems existing in the above-mentioned prior art. After the present invention performed MMBIT treatment on EH patients, it was found that after MMBIT treatment, the SLC9A1 protein and S100A2 protein in the blood of EH patients were significantly increased, and the purpose of improving the systolic and diastolic blood pressure of EH patients was finally achieved. It can be seen that SLC9A1 protein and S100A2 protein can be used as targets for the preparation of drugs for treating essential hypertension.

[0006] To achieve the above object, the present invention provides the following solutions:

[0007] The present invention provides the use of SLC9A1 protein and S100A2 protein as action targets in the preparation of a drug for treating essential hypertension.

[0008] Preferably, the effect of treating essential hypertension is exerted by increasing the expression levels of SLC9A1 protein and S100A2 protein.

[0009] As an additional solution, the present invention also provides the use of SLC9A1 protein and S100A2 protein as targets in the preparation of a Mongolian medicine for treating essential hypertension using the mind-body interaction therapy.

[0010] More preferably, the expression levels of SLC9A1 protein and S100A2 protein are increased to thereby achieve the effect of treating essential hypertension.

[0011] The present invention provides use of a SLC9A1 protein activator and a S100A2 protein activator in preparing a medicine for treating essential hypertension.

[0012] Preferably, the effect of treating essential hypertension is exerted by increasing the expression levels of SLC9A1 protein and S100A2 protein.

[0013] As an additional solution, the present invention also provides the use of SLC9A1 protein activator and S100A2 protein activator in the preparation of a Mongolian medicine for treating essential hypertension using the mind-body interaction therapy.

[0014] More preferably, the expression levels of SLC9A1 protein and S100A2 protein are increased to thereby achieve the effect of treating essential hypertension.

[0015] The present invention provides a medicine for treating essential hypertension, which comprises a SLC9A1 protein activator and a S100A2 protein activator.

[0016] Preferably, the drug plays a role in treating essential hypertension by increasing the expression levels of SLC9A1 protein and S100A2 protein.

[0017] Preferably, the drug further comprises pharmaceutically acceptable excipients.

[0018] As an additional scheme, the present invention also provides a medicine for treating primary hypertension using Mongolian medicine mind-body interaction therapy, wherein the medicine comprises a SLC9A1 protein activator and a S100A2 protein activator.

[0019] More preferably, the expression levels of SLC9A1 protein and S100A2 protein are increased to thereby achieve the effect of treating essential hypertension.

[0020] Further preferably, the medicine also includes pharmaceutically acceptable excipients.

[0021] The present invention provides the use of SLC9A1 protein and S100A2 protein as action targets in screening drugs for treating essential hypertension.

[0022] As an additional solution, the present invention also provides the use of SLC9A1 protein and S100A2 protein as targets in screening drugs for treating essential hypertension using Mongolian medicine mind-body interaction therapy.

[0023] The present invention provides a method for screening drugs for treating essential hypertension, by detecting the expression levels of SLC9A1 protein and S100A2 protein in subjects before and after drug administration.

[0024] As an additional scheme, the present invention also provides a method for screening drugs for treating essential hypertension using Mongolian medicine mind-body interaction therapy, by detecting the expression levels of SLC9A1 protein and S100A2 protein in the subjects before and after administration.

[0025] The present invention discloses the following technical effects:

[0026] The present invention provides the use of SLC9A1 protein and S100A2 protein as targets in the preparation of drugs for treating essential hypertension. In a specific embodiment of the present invention, MMBIT treatment was performed on EH patients, and the blood of EH patients before and after MMBIT treatment was analyzed. By comparing the proteome and RNA-seq, 187 differentially expressed genes and 56 differentially expressed proteins were identified, respectively; qPCR and transcriptome sequencing results were highly consistent; ELLSIA results showed that after MMBIT treatment, SLC9A1 protein and S100A2 protein in the plasma of EH patients were significantly higher than before MMBIT treatment (P < 0.05). In summary, increasing the expression of SLC9A1 protein and S100A2 protein in the plasma of EH patients can significantly improve the systolic and diastolic blood pressure of EH patients, that is, SLC9A1 protein and S100A2 protein can be used as targets for the preparation of drugs for treating essential hypertension.

[0027] MMBIT treatment can significantly increase the expression of SLC9A1 protein and S100A2 protein in the blood of EH patients. Therefore, the present invention uses MMBIT as a treatment method to treat EH patients again. The results once again verify that MMBIT treatment can significantly improve the blood pressure, somatic symptoms, anxiety and depression of EH patients. It can be seen that the SLC9A1 protein activator that increases the expression of SLC9A1 protein and the S100A2 protein activator that increases the expression of S100A2 protein can be used to treat EH. The present invention provides a new treatment approach for the treatment of EH. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0029] Figure 1 The protein sequencing results of plasma from different EH patients;

[0030] Figure 2 A volcano plot is drawn for the comparison group containing the T-test Pvalue value; the horizontal axis is the value of the Ratio value of the differential expression change of the comparison group after Log2 transformation; the vertical axis is the statistical test T-test and P-value after -Log 10 After conversion, red points indicate significantly up-regulated, blue points indicate significantly down-regulated, and gray indicates no significant difference; the names of up-regulated and down-regulated (Log2 Ratio absolute value from large to small) differential proteins are also marked;

[0031] Figure 3 The radar chart shows the top 30 differentially expressed proteins. The first circle is sorted clockwise along the orange arrow according to the absolute value of the difference after Log2 conversion from large to small. The second circle represents the Ratio value of Log2 conversion, pink represents upregulation, light blue represents downregulation, and the larger the point, the greater the difference. The third circle represents the average quantitative level of the two groups. If the expression level of the differentially expressed protein is high, a spike will appear in the chart.

[0032] Figure 4 It is a KEGG diagram; the horizontal axis is the number of differentially expressed proteins in the classification, and the vertical axis represents the secondary functional classification in the primary classification of KEGG (metabolism, genetic information processing, environmental information processing, cellular process, organism system and human disease); the primary classification of KEGG is represented by different colors in the figure;

[0033] Figure 5 This is a GO enrichment analysis diagram; GO Circos from outside to inside: the first circle is the enriched GO function, and different colors represent different categories; the second circle is the number of differentially expressed proteins and the significant P-value value contained in the function, and the color indicates the enrichment significant P-value value after -Log 10 The converted value, the larger the value, the redder the color; the third circle is a bar chart of the number of up- and down-regulated differential proteins, pink represents the number of up-regulated differential proteins, and blue represents the number of down-regulated differential proteins; the fourth circle is the enrichment degree of each function (Fold enrichment);

[0034] Figure 6 Comparative Venn diagram for transcriptome and proteome identification;

[0035] Figure 7 It is a scatter plot of transcripts and their corresponding protein expression levels; the horizontal axis is the protein expression level and the vertical axis is the transcript expression level. The color of the dot represents the density of the point at that location, and R represents the Pearson correlation coefficient.

[0036] Figure 8 Venn diagram for comparative analysis of differentially expressed proteins and differentially expressed transcripts;

[0037] Fig. 9 KEGG enrichment heat map; the horizontal axis is the different protein classifications, the vertical axis is the KEGG pathway, and the color of the heat map indicates the enrichment significance p-value through log 10 and Z score transformed values, the redder the color, the more significant the enrichment;

[0038] Fig.10 This is a heat map of biological process enrichment in GO. The horizontal axis is the different protein classifications, the vertical axis is the GO function, and the color of the heat map indicates the enrichment significance p-value through log 10 and Z score transformed values, the redder the color, the more significant the enrichment;

[0039] Fig.11 This is a heat map of cellular components in GO enrichment; the horizontal axis is different protein classifications, the vertical axis is GO function, and the color of the heat map indicates the enrichment significance p-value through log 10 and Z score transformed values, the redder the color, the more significant the enrichment;

[0040] Fig.12 Molecular function heat map of GO enrichment; the horizontal axis is different protein classifications, the vertical axis is GO function, and the color of the heat map indicates the enrichment significance p-value through log 10 and Z score transformed values, the redder the color, the more significant the enrichment;

[0041] Fig.13 The statistical results of the relative expression of SLC9A1 in the blood of EH patients before and after MMBIT treatment, n = 15; Pre-MMBIT refers to before MMBIT treatment, and Post-MMBIT refers to after MMBIT treatment;

[0042] Fig.14The statistical results of the relative expression of S100A2 in the blood of EH patients before and after MMBIT treatment, n = 20; Pre-MMBIT refers to before MMBIT treatment, and Post-MMBIT refers to after MMBIT treatment;

[0043] Fig.15 This is a diagram of the data screening process before and after MMBIT treatment;

[0044] Fig.16 This is a graph showing changes in blood pressure control rate;

[0045] Fig.17 This is the statistical result of the number of people whose scores are above the positive value and below the positive value of the SSS psychological scale. DETAILED DESCRIPTION

[0046] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0047] It should be understood that the terms described in the present invention are only for describing a particular embodiment and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0048] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0049] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to those skilled in the art. The present invention description and examples are exemplary only.

[0050] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0051] Example 1

[0052] 1. Research subjects

[0053] Clinical blood samples were collected from 25 patients with essential hypertension (EH patients) at the International Mongolian Medicine Hospital of Inner Mongolia Autonomous Region from January to December 2022. They needed to receive a 28-day course of MMBIT (Mongolian Mind-Body Interactive Treatment), with MMBIT treatment once a day. The experiment was approved by the ethics committee of the hospital (approval number: 2020-014).

[0054] Screening criteria: age 18-65 years old, regardless of gender, with a junior high school education or above; the diagnostic criteria for primary hypertension (diagnosed based on the "China Hypertension Prevention and Treatment Guidelines 2018 Revised Edition") are mild or moderate; the risk classification of primary hypertension is low risk; patients have not taken antihypertensive drugs since diagnosis; four infectious tests are negative; voluntarily participate in this clinical study and sign the informed consent.

[0055] 2. Experimental reagents

[0056] Human S100 calcium binding protein A2 (S100A2) ELISA kit (Jianglai Biotechnology, JL15077-48T); Human sodium / hydrogen exchange factor 1 (NHE1) ELISA kit (Jianglai Biotechnology, JL17896-48T); Trizon Reagent (CWBIO, CW0580S); Ultrapure RNA extraction kit (CWBIO, CW0581M); HiScript IIQ RT SuperMix for qPCR (+gDNAwiper) (Vazyme, R223-01); ChamQ Universal SYBR qPCR Master Mix (Vazyme, Q711-02); 50×TAE buffer (Solarbio, T1060); 6×DNA Loading Buffer (TRANS, GH101-01); 50bpDNALadder (TIANGEN, MD108); Gsafe Redplus nucleic acid dye (GLPBIO, GK20002); agarose powder (Invitrogen, 75510-019); MouseAnti-β-Actin (TransGen, HC201); RabbitAnti HTR2B (ABclonal, A5670).

[0057] 3. Instruments and Equipment

[0058] OMRON electronic sphygmomanometer (HEM-1000), LightCycler480 II fluorescence quantitative PCR instrument (Roche, Rotkreuz, Switzerland), TC-EAPCR amplifier (Hangzhou Bioray Technology Co., Ltd.), Chemi DocTM XRS+ ultra-high sensitivity chemiluminescence imaging system (Bio-Rad Biomedical Products (Shanghai) Co., Ltd.); proteome sequencing and analysis were commissioned to Jingjie Biotechnology Co., Ltd.; blood mRNA sequencing was commissioned to BGI Genomics Co., Ltd.

[0059] 4. Experimental methods

[0060] 4.1 Proteome sequencing and analysis

[0061] DIA quantitative proteome is based on magnetic bead protein sequencing, after protein extraction, trypsin hydrolysis, TMT labeling, HPLC fractionation and liquid chromatography-mass spectrometry analysis, when P value <0.05, the difference in expression changes more than 1.3 as the threshold of significant upregulation, less than 1 / 1.3 as the threshold of significant downregulation. For the annotation of protein pathways, the Kyoto Encyclopedia of Genes and Genomes (KEGG) database was used. Cluster analysis based on functional enrichment of different differential protein groups was used to study the potential associations and differences in their specific functions.

[0062] 4.2 Transcriptomic Sequencing and Analysis

[0063] Total RNA was extracted using an ultrapure RNA extraction kit (CWBIO, CW0581M), and quantified to construct an mRNA library. After sequence filtering and alignment, the FPKM value was calculated using Stringtie and TMM to characterize gene expression, and the differential expression fold (FC) was calculated. Differential genes were screened based on P value < 0.05 and FC > 1, and cluster analysis was performed using the Bioconductor package in R, and biological pathway enrichment analysis of differential genes was performed using KEGG and GO.

[0064] 4.3 Transcriptome and proteome association analysis

[0065] In order to explore the effect of MMBIT treatment on EH patients, the proteomic and transcriptomic expression data were jointly analyzed to analyze the potential relationship. The transcriptome and proteome are two different levels that reflect the essence of the organism. By comparing the quantitative correlation of the two groups, the potential regulatory relationship between proteins and transcripts can be quickly understood. Through the above protein and transcript ID correspondence, the protein expression and mRNA expression were merged.

[0066] 4.3.1. Annotation Methods

[0067] (1) Gene Ontology (GO) annotation

[0068] GO is an important bioinformatics initiative that aims to unify the representation of gene and gene product properties across all species. More specifically, the project aims to: 1. Maintain and develop a controlled vocabulary for gene and gene product properties; 2. Annotate genes and gene products, assimilate and disseminate annotation data; 3. Provide tools to easily access all aspects of the data provided by the project. The ontology covers three areas, as follows:

[0069] 1. Cellular component: A cellular component is a component of a cell, but only if it is part of a larger object; this may be an anatomical structure (such as the rough endoplasmic reticulum or the nucleus) or a group of gene products (such as ribosomes, proteasomes, or protein dimers); 2. Molecular function: Molecular function describes activities that occur at the molecular level, such as catalytic or binding activity; GO molecular function terms represent activities rather than entities (molecules or complexes) that perform actions, and do not specify the place, time, or context in which the actions occur; 3. Biological process: A biological process is a series of events that are accomplished by a combination of one or more ordered molecular functions; it can be difficult to distinguish between biological processes and molecular functions, but the general rule is that a process must have more than one distinct step.

[0070] The GO annotated protein group is derived from the UniProt-GOA database (http: / / www.ebi.ac.uk / GOA / ). First, the identified protein IDs are converted to UniProt IDs, and then mapped to GO IDs through protein IDs. If some of the identified proteins are not annotated by the UniProt-GOA database, InterProScan software is used to annotate the GO function of the protein based on the protein sequence alignment method, and then the protein is annotated with gene ontology based on the three major categories of biological process, cellular component and molecular function.

[0071] (2) KEGG pathway annotation

[0072] KEGG connects known molecular interaction network information, such as pathways and complexes (Pathway database), gene and protein information generated by genome projects (including gene databases), and biochemical compound and reaction information (including compound and reaction databases). These databases are different networks, called "protein networks" and "chemical universes", respectively. Efforts are currently underway to increase the knowledge of KEGG, including information about orthogonal clusters in the KEGG Orthology database. KEGG pathways mainly include: metabolism, genetic information processing, environmental information processing, cellular processes, human diseases, and drug development. The Kyoto Encyclopedia of Genes and Genomes (KEGG) database is used to annotate protein pathways, specifically: First, the KEGG database description of the protein is annotated using the KEGG online service tool KAAS, and then the annotation results are mapped to the KEGG pathway database using the KEGG online service tool KEGGmapper.

[0073] 4.3.2 Functional enrichment

[0074] (1) Enrichment of GO analysis

[0075] Proteins were divided into three categories: biological process, cellular component, and molecular function by GO annotation. For each category, a two-tailed Fisher's exact test was used to test the enrichment of differentially expressed proteins over all identified proteins. GO with a corrected p value < 0.05 was considered significant.

[0076] (2) Enrichment of pathway analysis

[0077] The KEGG database was used to identify enriched pathways, and the two-tailed Fisher's exact test was used to detect the enrichment of differentially expressed proteins for all identified proteins. Corrected p values ​​< 0.05 were considered significant pathways. According to the KEGG website, these pathways were divided into different hierarchical categories.

[0078] 4.3.3 Enrichment-based clustering

[0079] Further hierarchical clustering was performed based on the functional classification of differentially expressed proteins (e.g., GO, domain, pathway, complex). All categories and their P values ​​obtained after enrichment were sorted, and then the categories that were enriched in at least one of the clusters and had a P value < 0.05 were filtered. The filtered P value matrix was calculated by the function x = -log 10 (P value) transformation, and finally z-transformation of the x-values ​​for each functional category, and clustering of these z-scores using one-way hierarchical clustering (Euclidean distance, average linkage clustering). Cluster membership was visualized by heatmap using the “heatmap.2” function in the “gp lots” R-package.

[0080] 4.4. Detection of S100A2 and SLC9A1 in the blood of EH patients by ELSIA

[0081] Clinical blood samples were tested using human S100 calcium binding protein A2 (S100A2) ELISA kit and human sodium / hydrogen exchange factor 1 (NHE1) ELISA kit (perform the experiment according to the instructions).

[0082] 4.5 Data Analysis

[0083] Basic information data are presented as mean ± standard deviation (mean ± standard deviation). SPSS Statistics 17.0 was used for statistical analysis, and GraphPad Prism v8.0 was used for drawing. The inter-group comparison was statistically analyzed by chi-square test and nonparametric rank sum test according to the data, and p value < 0.05 was considered statistically significant.

[0084] 5. Results

[0085] 5.1. Blood pressure survey of EH patients before and after MMBIT treatment

[0086] In this example, 15 EH patients (9 females and 8 males) with an average age of 47.6±9.65 years old were surveyed. The blood pressure survey results before and after MMBIT treatment are shown in Table 1.

[0087] Table 1 Blood pressure survey of EH patients before and after MMBIT treatment

[0088] Before MMBIT treatment After MMBIT Treatment P Systolic blood pressure 149.0±14.89 142.3±13.10 0.0001** Diastolic blood pressure 88.73±11.72 85.69±8.94 0.0001**

[0089] Note: *: p < 0.05, **: p < 0.01, ns: no difference; Statistical method: Wilcoxon signed-rank test.

[0090] As shown in Table 1, both systolic and diastolic blood pressure of EH patients were significantly improved after MMBIT treatment.

[0091] 5.2 Proteome of EH patients before and after MMBIT treatment

[0092] Protein sequencing was performed on the plasma of 15 EH patients to detect the proteome of EH patients before and after MMBIT treatment. The results are as follows Figure 1-Figure 5 shown.

[0093] Compared with before MMBIT treatment, a total of 56 significant proteins were found before and after MMBIT treatment, involving 26 up-regulated proteins and 30 down-regulated proteins ( Figure 1); The volcano plot showed that the expression of FGR, CD63, MYH1, SLC9A1, and S100A2 was higher after treatment than before treatment, while the expression of PRCP, ECHDC1, PARP1, MBOAT2, etc. was lower after treatment than before treatment ( Figure 2 ). The radar chart of the top 30 differentially expressed proteins showed that ALYREF, CLNS, BET1, SCL9A1, ECHDC1, and PRCP were expressed at higher levels than the average quantitative level of the two ( Figure 3 Then, the 55 important proteins were analyzed for differential protein classification. In KEGG, there were more proteins in the cellular processes of cell migration and transport and catabolism. In the environmental information process, there were more proteins in signal transduction. In the human disease domain, there were more proteins in tumors and neruodegenerative disease. In the metabolic domain, there were more proteins in global and overview maps. In the organ system, there were more proteins in the endocrine system. Figure 4 ); in GO enrichment analysis, the molecular function analysis focused on protein phosphatase 2B binding and phosphatase activity; in cellular component analysis, the focus was on organelle envelope lumen, protein-DNA complex, and juxtaparanode region ofaxon in Cellular Comonent; in biological process analysis, the focus was on uronic acid metabolic process and myelin maintenance ( Figure 5 ).

[0094] 5.3 Correlation between proteome and transcriptome in EH patients before and after MMBIT treatment

[0095] The inventors performed high-throughput transcriptome sequencing on 6 EH patients before and after treatment, and screened out 92 differentially expressed genes, including 38 upregulated differentially expressed genes and 54 downregulated genes. Transcriptomics and proteomics describe the expression of genes at the transcriptional and translational levels, respectively. In addition to the one-to-one relationship between transcriptomes and proteomics, there are other more complex regulatory relationships. By comparing the quantitative correlation of the two omics, we can quickly understand the potential regulatory relationship between proteins and transcripts.

[0096] In order to count the intersection between the identified proteins and the identified mRNAs, the protein IDs were converted to the corresponding transcript IDs. According to the transcript IDs, the intersection of the two omics identifications was counted, and the number of transcript IDs was 14,847, the number of protein IDs was 2,897, and the number of intersection IDs was 2,644 ( Figure 6). From the scatter plot of transcripts and corresponding protein expression, we know that the two are positively correlated ( Figure 7 ). The Venn diagram revealed that there was an overlap in the up-regulated parts of the transcriptome and protein group, namely S100A2 ( Figure 8 ).

[0097] The p-value was used to compare the enrichment of these GO functions / KEGG pathways in different protein categories, and the GO functions / KEGG pathways that were significantly enriched (p<0.05) in at least one category were screened using a heat map. The results are shown in the figure below. Figure 9-12 shown.

[0098] The heat map visualization shows the different regulatory relationships between the transcriptome and the proteome. The results show that Necroptosis, Amyotrophic lateral scierosis, and mRNA surveillance pathway are downregulated in the KEGG enrichment set, while Gastric acid secretion, Bile secretion, Pancreatic secretion, and AMPK signaling pathway are upregulated. Fig. 9 ). Biological processes in GO enrichment up-regulate negative regulation of apoptotic process, sensory perception of sound, cardiac muscle contraction, etc., and down-regulate purineribonucleotide metabolic process, regulation of receptor-mediated endocytosis, etc. ( Fig.10 ); in the cellular component, the extracellular space, extracellular exosome, vesicle, etc. are upregulated, and the perinuclear region of cytoplasm, midbody, etc. are downregulated ( Fig.11 ); in Molecular function, it upregulates signaling receptor regulator activity, protein serine / threoninephosphatase activity, etc., and downregulates acyltransferase activity, structural constituent of cytoskeleton, etc. ( Fig.12 ).

[0099] 5.4. ELSIA test results

[0100] The statistical results of S100A2 and SLC9A1 levels in the blood of EH patients before and after MMBIT treatment are as follows Fig.13 and Fig.14 shown. Fig.13 The results showed that before MMBIT treatment, the relative expression of SLC9A1 in the blood of EH patients was 1.048±0.395, and after MMBIT treatment, the relative expression of SLC9A1 in the blood of EH patients was 1.953±1.09. Compared with before MMBIT treatment, the expression of SLC9A1 in the blood of EH patients increased significantly, with a significant difference (p=0.0003); Fig.14 The results showed that before MMBIT treatment, the relative expression of S100A2 in the blood of EH patients was 3.053±0.404, and after MMBIT treatment, the relative expression of S100A2 in the blood of EH patients was 4.1300±0.644. Compared with before MMBIT treatment, the expression of S100A2 in the blood of EH patients increased significantly, with a significant difference (p<0.0001).

[0101] Since MMBIT treatment can significantly increase the levels of S100A2 and SLC9A1 in the blood of EH patients, subsequent experiments treated EH patients by increasing the levels of S100A2 and SLC9A1, which once again verified that increasing the levels of S100A2 and SLC9A1 in the blood of EH patients can achieve the effect of treating EH.

[0102] Example 2

[0103] 1. Research subjects

[0104] The study subjects were EH patients who were able to cooperate with the questionnaire survey, that is, had a smartphone and could use it, were willing to participate in the survey, and received MMBIT for 21 days from June to July 2022. All patients voluntarily participated in the survey, and this study was approved by the Ethics Committee of the Sino-Mongolian Medical Research Institute of Inner Mongolia Autonomous Region (NO.2020-014).

[0105] 2. Questionnaire

[0106] The designed questionnaire was published on the questionnaire website platform. The questionnaire was published one week before and after the survey. Blood pressure measurement was performed at home using a home blood pressure monitor and uploaded to the questionnaire website in the form of pictures. The questionnaire consisted of 4 parts: (i) confirmation of consent and basic information characteristics of the respondents, (ii) somatization self-assessment scale, (iii) blood pressure measurement pictures, and (iv) satisfaction evaluation. Consent was implied by answering the questionnaire.

[0107] 2.1. Blood pressure judgment criteria

[0108] According to the 2018 "Revised Edition of the Guidelines for the Prevention and Treatment of Hypertension in China", a patient is diagnosed with EH if SBP ≥ 140 mmHg and / or DBP ≥ 90 mmHg; if the value is less than this after treatment, it is considered to be up to standard.

[0109] 2.2. Somatization Self-rating Scale (SSS)

[0110] SSS is a self-rating scale for somatic symptoms designed and compiled by Professor Mao Jialiang, a Chinese cardiologist. It consists of 20 questions: somatic symptom questions (S factor: cardiovascular symptoms: palpitations, chest tightness, chest pain, shortness of breath; sleep disorders: difficulty falling asleep, dreaming, easy awakening, early awakening, insomnia) account for 45%, anxiety (A factor) accounts for 25%, depression (D factor) accounts for 20%, and anxiety and depression (AD factor) accounts for 10%. Each question is divided into four levels according to the severity of the symptoms, namely none, mild, moderate and severe, and is scored as 1, 2, 3 and 4 points respectively. A total score greater than 36 points is considered positive.

[0111] 3. Data screening

[0112] In June 2022, the "Essential Hypertension Pre-treatment Assessment Scale" was released in the mobile app. 21 days later, the "Essential Hypertension Post-treatment Assessment Scale" was collected and data was screened. The data screening process is as follows: Fig.15 As shown. A total of 4484 samples were collected before MMBIT treatment. After excluding 37 samples diagnosed by non-tertiary hospitals, a preliminary screening was performed. Among the preliminary screening, 1263 blood pressure pictures did not meet the requirements (the reasons were: 39 pictures that were not uploaded, 33 mercury and mercury pictures, 354 drug pictures, and 837 other pictures), and 3184 results were obtained; then the 3184 results were rescreened, and 650 blood pressure pictures that did not meet the requirements were eliminated (the reasons were: 15 pictures that were not uploaded, 13 mercury and mercury pictures, 66 drug pictures, and 556 other pictures), leaving 2534; finally, the unmatched data before and after MMBIT treatment were eliminated, and finally 739 samples remained, that is, 739 valid questionnaires were filled out before and after MMBIT treatment, and then data quality control was performed on these 739 samples. The general information of the research subjects is shown in Table 2.

[0113] Table 2 General information of EH patients receiving MMBIT (n = 739)

[0114]

[0115] 4. Treatment methods

[0116] The online questionnaire was completed by 739 EH patients. All EH patients in this sample received 28 days of MMBIT treatment, which includes narrative therapy, cognitive therapy, mindfulness therapy, music therapy, catharsis therapy, health education and other comprehensive Mongolian localized mind-body treatment methods. The main process of this therapy includes: MMBIT is a group health treatment method that can treat 50-500 people at the same time depending on the treatment room. During the entire treatment process, about 6 patients talk about their diseases and symptoms, and how they overcome the symptoms after receiving MMBIT, and then experienced doctors make professional comments. The main process of video group therapy includes: Mongolian song introduction, treatment and therapist introduction, health exercises, case report and Mongolian song ending. The treatment course is 21 days per course; in the on-site treatment, the health exercises part is replaced by MMBIT theory, health and disease education, mind-body practice, diet and behavior therapy education and other related content. Both video group therapy and on-site treatment mainly use case report narrative therapy and hypnosis methods. It is recommended that EH patients participating in group therapy start with individual consultation. In addition, professional doctors also widely use psychoanalysis, cognitive, behavioral and supportive therapy for case analysis and commentary. Music is played before and after treatment to make EH patients feel comfortable and relieve stress. In MMBIT treatment, self-cognition and self-reflection of EH patients are important. Therefore, through the statements of other EH patients and the comments of professional doctors, EH patients can improve their cognition, let go of pain, increase self-reflection, and continue to deal with these problems. In MMBIT treatment, health education, case presentation and doctor's comments are used together to implement mental illness intervention.

[0117] 5. Statistical methods

[0118] SPSS13.0 software was used for statistical analysis. The measurement data were expressed as mean ± standard deviation (z ± s). The comparison was performed by t test. The difference was considered statistically significant when P < 0.05. The classification data were expressed as n (%). The comparison between groups was performed by X 2 Test. Parametric test was used for measurement data with normal distribution and homogeneous variance, and nonparametric test was used for data with nonnormal distribution and unequal variance. T test was used for intra-group comparison, and P ≤ 0.05 indicated statistically significant differences. The correlation between before and after treatment was calculated using Rcor package.

[0119] 6. Results

[0120] 6.1 Effects of MMBIT on blood pressure, physical symptoms and psychology in patients with EH

[0121] The results of EH patients who received 21 days of MMBIT showed that systolic blood pressure was significantly reduced after treatment (P≤0.001), while diastolic blood pressure had no statistically significant difference (P>0.05); somatic symptoms, anxiety, depression anxiety and the total of the somatization self-rating scale decreased significantly (p≤0.05), but there was no statistical difference in the depression factor (P>0.05) (as shown in Table 3).

[0122] Table 3 Comparison of physical symptoms, psychological distress and blood pressure before and after MMBIT treatment

[0123]

[0124] Note: n is the number of cases, and paired t-test was used for statistical analysis; there were 525 blood pressure images uploaded, so the number of cases for systolic and diastolic blood pressure was 525.

[0125] Changes in blood pressure control rate Fig.16 As shown in the results, the number of patients who achieved blood pressure control increased significantly after receiving MMBIT. In addition, the chi-square test using cross-tabulation revealed a significant difference in the blood pressure control rate before and after receiving MMBIT (X 2 =4.811; p=0.016).

[0126] The decrease in the positive value of the SSS psychological scale is as follows Fig.17 As shown in the figure, the number of people with positive values ​​on the SSS psychological scale decreased after MMBIT treatment, and there was a significant difference. Through the cross-table chi-square test, the positive rate after MMBIT treatment was significantly different from that before treatment (X 2 =5.727; p=0.010).

[0127] 6.2 Changes in the main outcomes of each subgroup

[0128] According to the basic information of the subjects, they were divided into subgroups, including gender, age, gender, education, duration of disease, whether they took antihypertensive drugs, whether they regularly tested blood pressure, whether they had hypertension in their immediate family, weekly exercise, whether they knew about hypertension prevention knowledge, etc. The results are shown in Table 4: According to whether they took antihypertensive drugs, the data before and after MMBIT treatment were compared. It was found that although the somatic symptoms of EH patients who did not take antihypertensive drugs were reduced, there was no significant difference; but anxiety was significantly reduced in both types of patients; and the systolic blood pressure of EH patients who took antihypertensive drugs was significantly reduced.

[0129] Table 4 Analysis of outcomes in each subgroup

[0130]

[0131] 6.3 Analysis of drug influencing factors on EH treatment effect

[0132] The data of EH patients who took antihypertensive drugs and those who did not were compared, and the results are shown in Table 5. The results showed that there were significant differences in clinical information such as age, education, course of disease, whether blood pressure was tested regularly, and whether there were family members with hypertension between the two groups.

[0133] Table 5 Differences in basic information characteristics of EH patients taking different antihypertensive drugs

[0134]

[0135]

[0136] The results of ordered classification regression analysis of different blood pressure treatment effects are shown in Table 6. The results show that whether or not to take antihypertensive drugs is not an independent factor in the effect of blood pressure treatment.

[0137] Table 6 Results of ordered classification regression analysis of different hypertension treatment effects

[0138] Classification p-value OR LL UL gender 0.507991211 1.1421001 0.770904737 1.694172337 age 0.752719832 1.064313268 0.7220185 1.568959468 Education level 0.669148692 1.085044977 0.746160024 1.578579257 Do you check your blood pressure regularly? 0.087211637 1.614039295 0.930882786 2.790794893 Whether taking antihypertensive drugs 0.735733138 0.929246612 0.606493907 1.422999528 Course of disease 0.190399815 0.909787129 0.78938939 1.04785319 Do you have any direct relatives with high blood pressure? 0.654155522 0.90537007 0.585672918 1.398490076 Weekly exercise 0.171356909 1.110827098 0.955712651 1.291938213 Do you know the knowledge about hypertension prevention? 0.636161393 0.792150829 0.302188984 2.078278514

[0139] EH patients with different disease courses were further compared, and the results are shown in Table 7. The results showed that only EH patients with a disease course of 1-5 years had significant changes in blood pressure and somatization scale. The patients in the group not taking antihypertensive drugs had a significant decrease in systolic blood pressure after 21 days of MMBIT treatment (P≤0.05), and the depression-anxiety factor in the somatization symptom scale had a significant decrease (P≤0.05). In EH patients taking antihypertensive drugs, systolic blood pressure, diastolic blood pressure, somatic factor, anxiety factor, depression-anxiety factor and total score were significantly reduced (P≤0.05). In the group of patients taking antihypertensive drugs, systolic blood pressure, diastolic blood pressure, somatic factor, anxiety factor, depression-anxiety factor and total score were significantly reduced (P≤0.05).

[0140] Table 7 MMBIT treatment of EH patients with a disease course of 1-5 years

[0141]

[0142]

[0143] Note: Wilcoxon test was used for statistical analysis, where * indicates p ≤ 0.05. This table only includes patients with a disease course of 1-5 years, a total of 230 patients.

[0144] Based on the results of the above embodiments, it can be seen that the present invention uses MMBIT as a psychological intervention method for EH patients. After 21 days of video or on-site treatment, the S100A2 and SLC9A1 levels in the blood of EH patients can be significantly increased, and finally the systolic blood pressure is significantly reduced from 136.07±18.85 mmHg before MMBIT treatment to 134.17±19.16 mmHg. At the same time, the present invention found that after MMBIT treatment, the somatization symptoms, anxiety, depression-anxiety and the total score of the somatization self-rating scale decreased significantly. The somatization symptom scale-somatization decreased from 18.76±5.65 to 17.71±5.8, the depression factor decreased from 7.69±2.61 to 7.54±2.7, the anxiety factor decreased from 7.33±2.75 to 6.89±2.67, the depression-anxiety factor decreased from 4.33±1.47 to 3.94±1.48, and the total score of the somatization symptom scale decreased from 38.12±11.38 to 36.08±11.75.

[0145] The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. Application of SLC9A1 protein and S100A2 protein as targets in the preparation of drugs for the treatment of essential hypertension.

2. The use according to claim 1, characterized in that: By increasing the expression of SLC9A1 protein and S100A2 protein, it plays a role in treating essential hypertension.

3. Application of SLC9A1 protein activator and S100A2 protein activator in the preparation of drugs for the treatment of essential hypertension.

4. The use according to claim 3, characterized in that: By increasing the expression of SLC9A1 protein and S100A2 protein, it plays a role in treating essential hypertension.

5. A drug for treating essential hypertension, characterized in that: The drugs include SLC9A1 protein activator and S100A2 protein activator.

6. The drug according to claim 5, characterized in that The drug plays a role in treating essential hypertension by increasing the expression levels of SLC9A1 protein and S100A2 protein.

7. The drug according to claim 5, characterized in that The medicine also includes pharmaceutically acceptable auxiliary materials.

8. Application of SLC9A1 protein and S100A2 protein as targets in screening drugs for the treatment of essential hypertension.

9. A method for screening drugs for treating essential hypertension, characterized in that: The expression levels of SLC9A1 protein and S100A2 protein in the subjects before and after administration were detected.

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