CN111518893A - Uremia marker and application thereof - Google Patents

Abstract

The invention provides a uremia marker and application thereof. The uremia marker comprises at least one of the following substances: ESRRG, TEAD2, HOXC10, TEAD1, FOS, GRHL1, PAX7, PAX3, PBX1, MAFG, GCM2, NR3C1, NR3C2, JUN, PBX2, SCRT2, MYBL2, SPDEF, STAT1, NEUROG2, JDP2, FOSB. The inventor finds that the expression levels of a plurality of transcription factors in samples of uremia patients and normal healthy control group populations have significant difference by means of the scATAC-seq technology, so that the transcription factors with different expression levels can be used as markers for diagnosis, prognosis evaluation or drug screening of uremia.

Description

Uremia marker and application thereof

Technical Field

The invention relates to the technical field of nephropathy, and particularly relates to a uremia marker and application thereof.

Background

Uremia (uremia) refers to a clinical syndrome consisting of a series of clinical manifestations and metabolic disorders that occur as a result of progressive decline in kidney function from various causes, through to loss of kidney function and eventually to the end stage. The disturbed internal environment is liable to cause the change of epigenetic characteristics of Chronic Kidney Disease (CKD) patients and influence gene expression through epigenetic mechanism, thereby accelerating the development of CKD to uremia (end stage kidney disease). This is also evidenced by the increasing number of studies at this stage. Studies have shown that CKD patients have varying degrees of DNA methylation, which is also affected by the uremic component. The inflammatory response in CKD5 stage patients leads to abnormal DNA hypermethylation and is associated with poor prognosis. When epigenetic detection is carried out on a renal tumor patient undergoing dialysis and a renal tumor patient with normal renal function, the fact that various genes of the former show hypermethylation of DNA proves that CKD and dialysis may promote hypermethylation of DNA. The above experimental results show that the uremic environment changes the epigenetic characteristics of the body, which in turn leads to the change of the transcription level. Unlike gene mutations, epigenetic changes are dynamic and possibly reversible, and thus, it is of great interest to analyze the development of disease from an epigenetic perspective.

ATAC-seq (Assay for Transposase-Access chromosome with high-throughput put sequencing) is an emerging scientific research technology for studying Chromatin openness (or Chromatin accessibility) from an epigenetic point of view based on high-throughput sequencing, and the principle is to utilize the characteristic that Transposase Tn5 is easy to specifically bind with open Chromatin, sequence DNA sequence fragments captured by Tn5 enzyme, compare the sequencing result with human genome information, analyze open regions of DNA, and predict Transcription Factor Binding Sites (TFBS) in the whole genome range through further motif analysis. Compared with the traditional DNase-Seq, FAIRE-Seq and other methods for researching open chromatin, the ATAC-Seq has the advantages of less cell quantity requirement, low cost, short period, large data, no need of antibody enrichment, capability of detecting the open state of chromatin in the whole genome range and the like. At present, the single-cell ATAC-Seq is widely applied to the fields of chromosome open map analysis, apparent modification difference research, embryonic development epigenetic modification, tumorigenesis epigenetic mechanism research, tumor heterogeneity and typing research, disease potential marker prediction and the like.

The mechanism of occurrence and development of uremia and its related complications is still unclear, and its specific biomarkers are not discovered at present. Therefore, there is a need to discover uremia-specific biomarkers by means of ATAC-seq technology.

Disclosure of Invention

The present invention is directed to solving at least one of the problems of the prior art. Therefore, the invention provides a uremia marker and application thereof.

In a first aspect, an embodiment of the present invention provides a marker for uremia comprising at least one of: ESRRG, TEAD2, HOXC10, TEAD1, FOS, GRHL1, PAX7, PAX3, PBX1, MAFG, GCM2, NR3C1, NR3C2, JUN, PBX2, SCRT2, MYBL2, SPDEF, STAT1, NEUROG2, JDP2, FOSB.

The intestinal flora marker provided by the embodiment of the invention at least has the following beneficial effects:

the inventor finds that the expression levels of a plurality of transcription factors in samples of uremia patients and normal healthy control group populations have significant difference by means of the scATAC-seq technology, so that the transcription factors with different expression levels can be used as markers for diagnosis, prognosis evaluation or drug screening of uremia.

In a second aspect, an embodiment of the present invention provides a use of the above-mentioned uremia marker in the preparation of a uremia diagnostic and/or prognostic reagent. The uremia marker is differentially expressed in uremia patients and normal healthy control groups, so that the diagnosis of uremia or the evaluation of the prognosis of uremia can be realized by quantitatively detecting the uremia marker in a sample of a subject.

In a third aspect, an embodiment of the present invention provides a use of the above-mentioned uremia marker in screening a drug for preventing and/or treating uremia. The uremia marker has differential expression in uremia patients and normal healthy control groups, so that the evaluation of the drug effect of candidate drugs can be realized by quantitatively detecting the uremia marker in a sample after drug administration, thereby achieving the purpose of screening drugs.

In a fourth aspect, an embodiment of the present invention provides a kit comprising reagents for quantitatively detecting the above-mentioned uremia marker. The reagent realizes the quantitative detection of the uremia marker in a sample of a subject, and after the reagent is compared with a preset threshold value, the obtained result can be effectively used for diagnosing uremia, or evaluating the prognosis of uremia, or screening uremia drugs.

In a fifth aspect, an embodiment of the present invention provides a use of a reagent for quantitatively detecting the above-mentioned uremia marker in preparing a diagnostic kit for uremia. The uremia diagnosis product obtained by the reagent can realize quantitative detection on the uremia marker in a sample of a subject, and the detection result can be accurately used for diagnosing uremia, evaluating the prognosis of uremia or screening uremia drugs after being compared with a preset threshold value.

According to some embodiments of the invention, the reagent quantitatively detects the uremic markers at the gene level and/or the protein level. For example, the content of nucleic acid fragments of the uremic marker in the sample is quantitatively determined by PCR, or the expression level of protein of the uremic marker in the sample is quantitatively determined by enzyme-linked immunoassay, radioimmunoassay, Western blotting, protein chip method, or the like.

In a sixth aspect, an embodiment of the present invention provides a method of assessing the risk of uremia, the method comprising the steps of:

(1) obtaining the relative abundance of the uremic markers in the subject sample;

(2) comparing the relative abundance with a preset threshold value, and judging the uremia risk according to the comparison result;

this method is not suitable for the diagnosis and/or treatment of diseases.

Wherein uremia risk refers to the risk of the subject suffering from uremia, or to the prognosis of the subject; or to the effect of a candidate drug on uremia. The preset threshold is a critical value of the uremia marker of the normal subject, and specifically can be a relative abundance value of the uremia marker measured from a normal healthy control group sample or a normalized relative abundance value.

According to the method of some embodiments of the present invention, when directly determining the risk of uremia according to the relative abundance result of the markers, at least one of the following conditions is satisfied, and it can be determined that the subject has uremia, or the prognosis of the uremia patient is poor, or the candidate drug does not meet the screening condition:

(1) the relative abundance of the ESRRG is higher than a preset threshold value;

(2) the relative abundance of TEAD2 is higher than a preset threshold;

(3) the relative abundance of HOXC10 is higher than a preset threshold;

(4) the relative abundance of TEAD1 is higher than a preset threshold;

(5) the relative abundance of FOS is higher than a preset threshold value;

(6) the relative abundance of GRHL1 is lower than a preset threshold value;

(7) the relative abundance of PAX7 is higher than a preset threshold;

(8) the relative abundance of PAX3 is higher than a preset threshold;

(9) the relative abundance of the PBX1 is higher than a preset threshold;

(10) the relative abundance of MAFG is higher than a preset threshold;

(11) the relative abundance of GCM2 is lower than a preset threshold;

(12) the relative abundance of NR3C1 is higher than a preset threshold;

(13) the relative abundance of NR3C2 is higher than a preset threshold;

(14) the relative abundance of JUN is lower than a preset threshold value;

(15) the relative abundance of PBX2 is lower than a preset threshold;

(16) the relative abundance of the SCRT2 is lower than a preset threshold;

(17) the relative abundance of MYBL2 is lower than a preset threshold;

(18) the relative abundance of the SPDEF is lower than a preset threshold;

(19) the relative abundance of STAT1 is higher than a preset threshold;

(20) relative abundance of NEUROG2 is higher than a preset threshold;

(21) the relative abundance of JDP2 is below a preset threshold;

(22) the relative abundance of FOSBs is below a predetermined threshold.

In a seventh aspect, an embodiment of the present invention provides a computer-readable storage medium storing computer-executable instructions for performing the above-mentioned method. By performing the above method, it is possible to accurately and efficiently determine whether a subject has uremia, or the prognosis of a uremia patient, or the uremia efficacy on a candidate drug.

In an eighth aspect, an embodiment of the present invention provides a system for assessing risk of uremia, the system comprising:

a detection device for determining the relative abundance of a uremic marker in a sample from a subject;

and the comparison device is used for comparing the relative abundance with a preset threshold value and judging the uremia risk according to a comparison result.

Drawings

FIG. 1 shows the results of differential transcription factor motif identification of uremic PBMC subpopulations according to example 1 of the present invention.

FIG. 2 shows the results of the transcription factor function enrichment and inflammatory response analysis of the uremic patient of example 1 of the present invention.

Fig. 3 is a schematic composition diagram of a system for assessing risk of uremia according to example 2 of the present invention.

Detailed Description

The concept and technical effects of the present invention will be clearly and completely described below in conjunction with the embodiments to fully understand the objects, features and effects of the present invention. It is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments, and those skilled in the art can obtain other embodiments without inventive effort based on the embodiments of the present invention, and all embodiments are within the protection scope of the present invention.

Example 1

1. Selection of the subject

20 samples were selected, and were each a disease group: 10 patients in uremic phase; normal healthy control group: 10 healthy control volunteers. All uremia patients are patients from hemodialysis centers of people's hospitals in Shenzhen city, chronic nephritis is caused, and the disease history of hypertension, diabetes, secondary nephropathy (such as systemic lupus erythematosus) and the like is avoided. All volunteers in the healthy control group are from physical examination department of people hospital in Shenzhen city, and the healthy control group and the disease group are matched in race, age and gender. During the study, all uremic patients and healthy control volunteers were not administered lipid-lowering drugs, hormones, and immunosuppressants.

2. Acquisition of Peripheral Blood Mononuclear Cell (PBMC) samples

(1) 3 mL/person of peripheral venous blood of a study object with an empty stomach of more than 8h is extracted, diluted by physiological saline with equal volume, mixed evenly and added into 6mL of lymphocyte separation liquid.

(2) Centrifuging at 2000rpm and 20 deg.C for 15 min; the visible liquid level after centrifugation is divided from bottom to top: the erythrocyte layer, the lymph layer, the mononuclear cell layer and the plasma layer are collected and transferred into a sterile centrifuge tube.

(3) PBS was added, centrifuged at 2000rpm for 15min at 20 ℃ and the supernatant removed and repeated once.

(4) Adding 1mL erythrocyte lysate, mixing, transferring to 1.5mL sterile EP tube, standing at 4 deg.C for 40min, centrifuging at 1600rpm and 20 deg.C for 5min, and discarding supernatant to obtain cell precipitate.

(5) Adding 1ml PBS, centrifuging at 1600rpm and 20 deg.C for 5min, discarding supernatant to obtain cell precipitate, and repeating the operation twice.

(6) PBMCs from 10 uremic patients and 10 control groups were mixed with PBS containing 0.04% BSA and concentrated to 1mL each to prepare PBMC suspensions. Storing in a refrigerator at 4 deg.C.

Scata-seq PBMC suspension treatment and sequencing on machine

(1) 20 μ L of PBMC suspension and 20 μ L of 2% Trypan blue were mixed well and 20 μ L of the mixture was pipetted from the mixture and added to a hemocytometer plate for counting. PBMC were diluted to 2 with 0.04% BSA in PBS×10⁶one/mL and filtered using a 40 μm Flowmi cell filter. The cell viability of the viable filtered PBMC calculated using trypan blue was greater than 85% for further experiments.

(2) Transfer 1 × 10⁶The PBMC was centrifuged at 3000g for 5min at 20 ℃ in a sterile centrifuge tube and the supernatant discarded. Adding 100 μ L of cold lysis buffer, mixing and ice-bath for 3 min; adding 1mL of cold cleaning solution, centrifuging at 20 ℃ for 5min by 500g, and removing the supernatant; adding cold cell nucleus buffer solution diluted by 10 times, and storing in ice bath for later use.

(3) Nuclei were counted using a blood cell counting plate and a microscope. Mixing 10 μ L of 30% ATAC transposase and 5 μ L of diluted cell nucleus solution, incubating at 37 deg.C for 60min, and storing in 4 deg.C refrigerator. Mixing 15 μ L of the above cell nucleus solution and 65 μ L of the reaction mixture, taking 75 μ L of the above mixed sample and gel beads to 10 Xgenomics chromium chip (10 Xgenomics, PN-1000086), and forming GEMs wrapped by oil drops by using microfluidic technology; mu.L of GEMs were pipetted from the recovery well into a PCR-8 rack tube, and the gel beads in the GEMs were dissolved using a thermal cycler to generate 10 XBarode-labeled single-stranded DNA.

(4) Add 125. mu.L of recovery reagent to each sample, mix well and centrifuge, remove the bottom 125. mu.L of red oil layer. Add 200. mu.L of the cleaning mixture, mix well with shaking, then stand at room temperature for 10min, remove the supernatant. The precipitate was washed with 300. mu.L of 80% ethanol, and after standing for 30s, the ethanol was removed. Adding 80% ethanol 200 μ L, standing for 30s, centrifuging to remove ethanol, adding eluent I40.5 μ L, mixing, standing at room temperature for 1min, clarifying solution, and transferring 40 μ L sample to new calandria. The sample was purified using the SPRIselect Reagent Kit, the purified sample was mixed with 48 μ L of SPRIselect Reagent, reacted at room temperature for 5min, and the supernatant was removed after centrifugation. Adding 80% ethanol 200 μ L, standing for 30s, removing ethanol, and washing repeatedly. Adding 40.5 mu L of Buffer EB, uniformly mixing, standing for 2min at room temperature, clarifying the solution, and transferring 40 mu L of sample to a new calandria; storage at-20 deg.C (2 weeks).

(5) The sample index amplification reagents and 40. mu.L of sample were mixed and the sample was amplified using a thermal cycler. The sample was mixed with 40 μ LSPRIselect Reagent and after 5min of reaction 130 μ L of supernatant was transferred to a new calandria. The sample was mixed with 74. mu.L of LSPRIselect Reagent, reacted at room temperature for 5min and the supernatant removed. Add 80% ethanol 200. mu.L, after standing for 30s remove ethanol and repeat the washing once. Adding 20.5 μ L Buffer EB, mixing, standing for 2min, transferring 20 μ L clarified sample to new calandria, and storing in refrigerator at-20 deg.C. The distribution of the library was evaluated by measuring the concentration of the library sample using Qubit3.0 (reference concentration for DNA concentration detection interval: 0.2-100 ng/. mu.L), measuring 1. mu.L of the sample on Agilent 2100 chip, and determining the size of the DNA fragment. Using a library Quantification Kit (KAPALibrary Quantification Kit, KAPAbiosystems, KK4824) to detect DNA concentrations ranging from 150-1000kb, samples qualified for quality testing were screened for subsequent sequencing on the Illumina platform.

Bioinformatics analysis of scATAC-seq data

Preprocessing raw data: firstly, filtering a sequence with a connector by using Trimmomatic; removing sequences containing > 5% of indeterminate base information; low quality sequences are filtered (the number of bases with a quality value of <10 is more than 20% of the total sequence).

And (3) data comparison: the effective sequences screened after the above treatment were aligned using bowtie with reference to human genome GRCh38 to analyze the distribution of DNA sequences on the genome.

Analyzing data:

1) enrichment peak (peak) identification: peak identification was performed using MACS software (peak harvesting) with default P <10-5, identifying ATAC-rich peaks on the genome.

2) Peak length distribution: the DNA fragment length of the open region is identified according to ATAC-seq, and the Count of the enrichment peak is counted.

3) Analyzing the enrichment condition of TSS (transcription Start site), the value of FRiP (fraction of read in captured Peak) to evaluate the enrichment effect of the sample, and calculating the value of IDR (reproduction Discovery Rate) to evaluate the consistency of peak among repeated samples.

4) Peak-related gene annotation: genes of the open chromatin regions were searched and, based on the annotation information, a ratio map of enrichment peaks over different genomic features was drawn.

5) Different cell subsets were identified by PCA analysis, which allowed the classification of phenotypically very similar cells.

6) And (3) analyzing the difference peak among samples, comparing the difference of the opening degree of the chromatin of different cells in different periods, and revealing the specific epigenetic characteristic of the environmental chromatin of the uremia.

7) According to the enrichment of the transcription factor motif, a transcription initiation site and related transcription factors are analyzed, and the transcription factors which are key to uremia are explored.

8) Analyzing related genes of the difference peak, and carrying out GO and KEGG enrichment analysis on the related genes, analyzing the biological function change caused by the difference genes, and predicting the relevance of the biological function change and the uremia complications.

RNA-seq PBMC suspension treatment and sequencing on machine

(1) The cells obtained by the operation of step 3 were centrifuged (about 1 × 10⁶Tube/tube), discard the supernatant and add 1mL of PBS, gently suspend the cell pellet, and transfer to a 2mL spin-top conical centrifuge tube. Centrifuging to remove PBS, adding 1mL cell lysate, repeatedly blowing to obtain clear and non-viscous liquid, standing at room temperature for 5min, and storing in refrigerator at-80 deg.C.

(2) Thawing the above liquid at room temperature, adding chloroform 200 μ L, mixing, standing on ice for 3min, 12000rpm, and centrifuging at 4 deg.C for 5 min. Transferring the upper water phase to a new EP tube, adding equivalent isopropanol, mixing uniformly, performing ice bath for 10min, centrifuging, removing supernatant, adding 600 μ L of 75% ethanol, mixing uniformly, centrifuging, removing supernatant, drying at room temperature for 10min, and adding 50 μ L of pure water without RNase to dissolve precipitate. The quality of the extracted RNA was assessed by spectrophotometry and capillary electrophoresis in combination with the RNA6000Nano kit.

(3) mRNA was enriched and purified using magnetic beads (with polyA tail). The treated mRNA was fragmented by dephosphorylation and phosphate addition treatment, so that a phosphate group was added to the 5 'end of the mRNA and a hydroxyl group was added to the 3' end. T4RNA ligase 2 was used to ligate an adenylated single stranded DNA3 'linker and a 5' linker to each end of the mRNA. cDNA was synthesized by reverse transcription. The RT products were screened for 350-450 base length using brads. The data library was constructed after 15 cycles of PCR amplification. The Illumina Hiseq 2500 was used for sequencing with a sequencing read length of 2 × 150bp paired end.

Bioinformatic analysis of RNA-seq data

And (3) screening data: firstly, filtering a sequence with a joint; sequences containing > 5% of indeterminate base information were removed and low quality sequences were filtered. The alignment was performed using the TopHat2 program with reference to the human genome GRCh38, and the number of sequences, distribution of regions, etc. of the alignment of the sequencing data with GRCh38 were analyzed.

And (3) data analysis:

1) analysis of Gene expression level: the level of expression of each gene was analyzed using FPKM 2;

2) performing gene differential expression analysis, performing differential analysis on genes in different groups, wherein the differential threshold value is P less than 0.05, and screening candidate genes with significant changes (3-5 times) for further analysis;

3) differential gene function analysis, GO enrichment analysis and KEGG signal path enrichment analysis.

The chromosome open region of each cell subtype in the uremia environment is analyzed by combining the scataC-seq data and the RNA-seq data, a potential regulation and control original catalog and related transcription factors are determined, a transcription and control network of PMBC in the uremia environment is described, and the key regulation and control network of uremia occurrence and development is analyzed by combining clinical data.

Results are shown in FIG. 1, and FIG. 1 shows the results of identifying the differential transcription factor motif of uremic PBMC subpopulation of example 1 of the present invention. Wherein A, B, C, D, E represents the identification results of differential transcription factor motif of uremic patients in CD4+ T cell subset, CD8+ T cell subset, T helper 17cell (Thelper 17cells) subset, NK cell subset and B cell subset of PBMCs, respectively. The left column in the diagram is a volcano diagram for cluster analysis, and the left side of the abscissa-0.1 in the volcano diagram is down-regulated expression and the right side of 0.1 in the volcano diagram is up-regulated expression; the middle column is a heat map of clustering analysis, NC _ PBMC in the heat map is a normal healthy control group, UR _ PBMC is a uremia patient group; the right column is the base profile of motif.

As can be seen from fig. 1, in contrast to the normal healthy control group, in the uremic environment:

(1) in CD4+ T cells, TEAD 2motif, HOXC10 motif, TEAD1 motif and FOS motif are up-regulated and GRHL1 motif is down-regulated;

(2) in CD8+ T cells, PAX7 motif, PAX3 motif, PBX1 motif and MAFG motif are up-regulated to express, and GCM 2motif, MYBL 2motif and NFYA motif are down-regulated to express;

(3) in T helper 17cells, STAT1 motif, NEUROG 2motif and TEAD 2motif are up-regulated and JDP2motif and FOSB motif are down-regulated;

(4) in NK cells, TEAD1 motif, TEAD 2motif, NR3C1 motif and NR3C 2motif are up-regulated and JUN motif, PBX 2motif, SCRT 2motif, MYBL 2motif and SPDEF motif are down-regulated;

(5) in B cells, the expression of ESRRG motif is up-regulated.

Because of the specificity of the binding site sequence of the transcription factor when the transcription factor is combined with the DNA sequence, the differential expression of the transcription factor motif can reflect that the transcription factor aiming at the binding site sequence also has obvious differential expression in uremia patients. Thus, the differential transcription factor identified above can be used for diagnosis, prognosis or screening of uremia.

FIG. 2 shows the results of the transcription factor function enrichment and inflammatory response analysis of uremic patients according to an embodiment of the present invention. Wherein, A is GO enrichment analysis result, B and C are KEGG enrichment analysis result, and D is inflammation reaction analysis result. As can be seen from the figure, the identified differential transcription factors activate the body's inflammation and immune-related pathways to varying degrees: TNF signaling pathway, P53 signaling pathway, Wnt signaling pathway, IL-17 inflammatory pathway.

Example 2

The present embodiments provide a system for assessing the risk of uremia. Fig. 3 is a schematic composition diagram of a system for assessing risk of uremia according to example 2 of the present invention. Referring to fig. 3, the present embodiment provides a system for assessing risk of uremia, the system comprising: a nucleic acid isolation apparatus 100 for isolating a nucleic acid sample from a stool sample of a subject; and the sequencing device 110 is used for detecting the uremia marker in the nucleic acid sample, comparing the relative abundance of the marker with a preset threshold value, and judging the uremia risk according to the relative size of the marker and the preset threshold value.

Example 3

The present embodiments provide a computer-readable storage medium having stored thereon computer-executable instructions for performing a method comprising:

(1) obtaining the relative abundance of the uremic markers in the subject sample;

(2) comparing the relative abundance with a preset threshold, judging whether the subject has uremia or not according to the comparison result, judging the prognosis condition of the uremia patient, or screening the medicine.

Example 4

The present embodiment provides a protein chip, which is coated with antibodies against ESRRG, TEAD2, TEAD1, FOS, and MAFG, and the protein chip and corresponding detection reagents can be used to detect the relative abundance of the above markers in a serum sample of a subject, and the detection result is compared with a preset threshold, so as to effectively determine whether the subject has uremia.

The embodiments of the present invention have been described in detail with reference to the accompanying drawings, but the present invention is not limited to the above embodiments, and various changes can be made within the knowledge of those skilled in the art without departing from the gist of the present invention. Furthermore, the embodiments of the present invention and the features of the embodiments may be combined with each other without conflict.

Claims (9)

1. A uremic marker comprising at least one of: ESRRG, TEAD2, HOXC10, TEAD1, FOS, GRHL1, PAX7, PAX3, PBX1, MAFG, GCM2, NR3C1, NR3C2, JUN, PBX2, SCRT2, MYBL2, SPDEF, STAT1, NEUROG2, JDP2, FOSB.

2. Use of the uremic marker according to claim 1 for the preparation of a uremic diagnostic and/or prognostic reagent.

3. Use of the uremic marker of claim 1 for screening a medicament for the prevention and/or treatment of uremia.

4. A kit comprising reagents for quantitatively detecting the uremic marker of claim 1.

5. Use of a reagent for quantitatively detecting the uremic marker of claim 1 in the preparation of a diagnostic kit for uremia.

6. The use according to claim 5, wherein the reagent quantitatively detects the uremic marker according to claim 1 at the gene level and/or protein level.

7. A method of assessing the risk of uremia comprising the steps of:

(1) obtaining a relative abundance of the uremic marker of claim 1 in a sample from a subject;

(2) comparing the relative abundance with a preset threshold value, and judging the uremia risk according to a comparison result;

the method is not suitable for the diagnosis or treatment of diseases.

8. A computer-readable storage medium having stored thereon computer-executable instructions for performing the method of claim 7.

9. A system for assessing the risk of uremia, comprising:

a detection device for determining the relative abundance of the uremic marker of claim 1 in a sample from a subject;

and the comparison device is used for comparing the relative abundance with a preset threshold value and judging the uremia risk according to a comparison result.

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