CN112852950A

CN112852950A - Acute myocardial infarction biomarker and application thereof

Info

Publication number: CN112852950A
Application number: CN202110203460.4A
Authority: CN
Inventors: 张步春; 马礼坤
Original assignee: Individual
Current assignee: Anhui Provincial Hospital First Affiliated Hospital of USTC
Priority date: 2021-02-23
Filing date: 2021-02-23
Publication date: 2021-05-28
Anticipated expiration: 2041-02-23
Also published as: CN112852950B

Abstract

The invention belongs to the field of myocardial infarction diagnosis research, and provides application of miR-532-5p and/or miR-766-5p in preparation of acute myocardial infarction diagnosis products. According to the invention, the miR-532-5p and/or miR-766-5p from serum exosomes can be used as a diagnosis marker for diagnosing acute myocardial infarction for the first time, and the miR-532-5p and/or miR-766-5p from serum exosomes is used as a marker for diagnosing acute myocardial infarction or prognostically judging acute myocardial infarction, so that the acute myocardial infarction is more accurate and rapid to diagnose, and has high specificity and good sensitivity.

Description

Acute myocardial infarction biomarker and application thereof

Technical Field

The invention belongs to the field of myocardial infarction diagnosis research, and particularly relates to an acute myocardial infarction biomarker and application thereof.

Background

Acute Myocardial Infarction (AMI) is myocardial necrosis caused by acute, persistent ischemia and hypoxia of coronary arteries. Clinically, severe and persistent poststernal pain, rest and incomplete relief of nitrate medicines are caused, and the increased activity of serum myocardial enzyme and progressive electrocardiogram change are accompanied, so that arrhythmia, shock or heart failure can occur, and the life can be threatened.

Methods for diagnosing acute myocardial infarction in the prior art include: (1) the electrocardiogram characteristic changes into new Q wave and ST segment elevation and ST-T dynamic evolution; (2) serum biomarkers of myocardial necrosis are elevated, creatine kinase isoenzyme (CK-MB) and troponin (T or I) are elevated. The disease can be increased after 3-6 hours of attack, CK-MB can be recovered to be normal after 3-4 days, and troponin can be recovered to be normal after 11-14 days. Regarding the diagnostic method for characteristic changes of electrocardiogram, due to the lack of clinical control studies of larger samples, the sensitivity and specificity of the current electrocardiogram diagnostic standard of AMI recommended by various authorities are not clear, and no unified standard exists, so that the application of the standard is limited. The specificity and accuracy of detection of one serum protein marker are not satisfactory.

Disclosure of Invention

In order to overcome the problems in the prior art, the invention aims to provide an acute myocardial infarction biomarker and application thereof.

In order to achieve the above objects and other related objects, the present invention adopts the following technical solutions:

in the first aspect of the invention, the application of miR-532-5p and/or miR-766-5p from serum exosomes in preparing acute myocardial infarction diagnosis products is provided.

In the second aspect of the invention, the application of the reagent for specifically recognizing miR-532-5p and/or miR-766-5p in preparing an acute myocardial infarction diagnosis kit is provided.

In a third aspect of the invention, an acute myocardial infarction detection kit is provided, and the kit at least comprises an acute myocardial infarction detection reagent, wherein the acute myocardial infarction detection reagent is selected from a reagent for specifically recognizing miR-532-5p and/or miR-766-5 p.

Compared with the prior art, the invention has the following beneficial effects:

according to the invention, miR-532-5p and/or miR-766-5p from a serum exosome is found to be a diagnosis marker for diagnosing acute myocardial infarction for the first time, and miR-532-5p and/or miR-766-5p is used as a marker for diagnosing acute myocardial infarction or prognostically judging acute myocardial infarction, so that the acute myocardial infarction is more accurate and rapid to diagnose, and has high specificity and good sensitivity.

Drawings

FIG. 1 identification and analysis of exosomes in serum (A transmission electron microscopy; B. Nanoparticle Tracking Analysis (NTA) assay).

Figure 2A volcano plots of serum exosomes differentially expressing mirnas (red for up-regulation and green for down-regulation).

Figure 2B hierarchical cluster analysis of serum exosomes differentially expressing mirnas (red for up-fold, blue for down-fold).

Figure 3 qRT-PCR verifies the relative expression levels of the miRNAs differentially expressed by the first 5 plasma exosomes of fold expression.

FIG. 4A diagnostic potency-ROC plot of plasma exosome-derived hsa-miR-766-5p for acute ST-elevated myocardial infarction (STEMI).

FIG. 4B diagnostic potency-ROC plot of plasma exosome-derived hsa-miR-532-5p for acute ST-elevated myocardial infarction (STEMI).

Figure 4C diagnostic potency-ROC graph of serum blood hypersensitivity troponin for acute ST elevation myocardial infarction (STEMI).

FIG. 5 KEGG pathway analysis of exosome differentially expressed miRNAs target genes.

Detailed Description

When numerical ranges are given in the examples, it is understood that both endpoints of each of the numerical ranges and any value therebetween can be selected unless the invention otherwise indicated. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In addition to the specific methods, devices, and materials used in the examples, any methods, devices, and materials similar or equivalent to those described in the examples may be used in the practice of the invention in addition to the specific methods, devices, and materials used in the examples, in keeping with the knowledge of one skilled in the art and with the description of the invention.

Unless otherwise indicated, the experimental methods, detection methods, and preparation methods disclosed herein all employ techniques conventional in the art of molecular biology, biochemistry, chromatin structure and analysis, analytical chemistry, cell culture, recombinant DNA technology, and related arts.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. In addition, any methods and materials similar or equivalent to those described herein can be used in the practice of the present invention. The preferred embodiments and materials described herein are intended to be exemplary only.

One embodiment of the invention relates to application of miR-532-5p and/or miR-766-5p from serum exosomes in preparation of acute myocardial infarction diagnosis products.

Specifically, miR-532-5p and/or miR-766-5p from serum exosomes is used as a biomarker.

The Access number of the miR-532-5p is as follows: NR _ 030241.2.

The Access number of the miR-766-5p is as follows: LM 383020.1.

The acute myocardial infarction diagnosis product is used for judging acute myocardial infarction, selecting a treatment scheme and/or evaluating prognosis;

the prognosis evaluation of acute myocardial infarction refers to the prognosis judgment of the course and/or the fate of a patient with acute myocardial infarction.

The acute myocardial infarction diagnosis product comprises an acute myocardial infarction detection reagent, wherein the acute myocardial infarction detection reagent is a reagent for specifically identifying miR-532-5p and/or miR-766-5 p.

It should be noted that the acute myocardial infarction detection reagent is not limited to necessarily be in a liquid form.

Further, the acute myocardial infarction detection reagent refers to a reagent for detecting the expression quantity of miR-532-5 and/or miR-766-5p in a sample, and whether a patient suffers from acute myocardial infarction, which treatment scheme is selected and/or the course and/or outcome of the patient are/is subjected to prognosis judgment according to the detection result.

The sample may be a plasma RNA sample of a subject to be tested.

The miR-532-5p and/or miR-766-5p is originated from exosomes of a patient.

In one embodiment, the agent that specifically recognizes miR-532-5p and/or miR-766-5p is selected from any one or more of the following:

1) a primer pair for specifically amplifying miR-532-5p and/or miR-766-5 p;

2) and the probe specifically recognizes the miR-532-5p and/or miR-766-5 p.

In one embodiment, when the reagent specifically recognizing miR-532-5p and/or miR-766-5p is selected from a primer pair specifically amplifying miR-532-5p and/or miR-766-5p, the primer pair specifically amplifying miR-532-5p and/or miR-766-5p comprises:

as shown in SEQ ID NO: 1, specifically amplifying an upstream primer sequence of miR-532-5 p; specifically, CCTTGAGTGTAGGACCGTAA.

As shown in SEQ ID NO: 2, a downstream primer sequence of the specific amplification miR-532-5 p; specifically, CATGCCTTGAGTGTAGGACCGT.

As shown in SEQ ID NO: 3, and specifically amplifying an upstream primer sequence of miR-766-5 p; specifically, AGGAATTGGTGCTGGTCTTAA.

As shown in SEQ ID NO: 4, and specifically amplifying a downstream primer sequence of the miR-766-5 p. Specifically, AGGAGGAATTGGTGCTGGTCTT.

The embodiment of the invention provides application of a reagent for specifically identifying miR-532-5p and/or miR-766-5p in preparation of an acute myocardial infarction diagnosis kit.

The reagent for specifically recognizing miR-532-5p and/or miR-766-5p is selected from any one or more of the following reagents:

(1) a primer pair for specifically amplifying miR-532-5p and/or miR-766-5 p;

(2) and the probe specifically recognizes the miR-532-5p and/or miR-766-5 p.

as shown in SEQ ID NO: 1, specifically amplifying an upstream primer sequence of miR-532-5 p;

as shown in SEQ ID NO: 2, a downstream primer sequence of the specific amplification miR-532-5 p;

as shown in SEQ ID NO: 3, and specifically amplifying an upstream primer sequence of miR-766-5 p;

as shown in SEQ ID NO: 4, and specifically amplifying a downstream primer sequence of the miR-766-5 p.

The acute myocardial infarction detection kit provided by the embodiment of the invention at least comprises an acute myocardial infarction detection reagent, wherein the acute myocardial infarction detection reagent is selected from a reagent for specifically recognizing miR-532-5p and/or miR-766-5 p.

(1) a primer pair for specifically amplifying miR-532-5p and/or miR-766-5 p;

(2) and the probe specifically recognizes the miR-532-5p and/or miR-766-5 p.

The kit can be a real-time fluorescent quantitative PCR detection kit, and the basic principle of the kit is that a pair of specific primers of target polynucleotide is utilized, and in the PCR amplification reaction solution of the kit, the circular amplification of the target polynucleotide is realized through a fluorescent quantitative PCR amplification instrument, so that the aim of quickly and quantitatively detecting the polynucleotide is fulfilled.

The present invention is not particularly limited to other components and final concentrations thereof in the PCR reaction system except for using the primer of the present invention, and one skilled in the art can establish the PCR reaction system based on the general components and concentrations thereof conventionally used in establishing the PCR system. The template (e.g., genomic DNA) for PCR amplification can also be extracted using methods conventional in the art.

The kit is used for detecting by adopting a real-time fluorescent quantitative PCR technology, and other conventional reagents required by PCR can be further included in the kit. Since the common PCR reagents can be purchased separately or configured by themselves through the market, the reagents can be assembled into the kit according to the actual needs of customers, and can be assembled into the kit for convenience.

The kit of the present invention may contain each set of primer pairs packaged independently, or may contain a prepared PCR detection mixture containing each set of primer pairs.

Example 1

Subject selection:

and (3) inclusion standard: 1) the acute ST elevation myocardial infarction (STEMI) patient group is incorporated into the standard reference 2012 global uniform definition of myocardial infarction: serum cardiac markers, mainly troponin, are elevated, at least exceed 99% of the reference value upper limit, accompanied by CAG or CCTA to confirm the formation of thrombus in coronary artery, one or more coronary artery main trunk or branch stenosis, the degree is greater than 70%; the ST segment of the electrocardiogram is lifted upwards from the dorsum of the arch.

(2) Healthy control groups were included as standards: the sex is the same as that of the experimental group, the electrocardiogram has no coronary heart disease characteristic, the serum myocardial enzyme marker is mainly normal troponin, and CAG or CCTA shows that no abnormal stenosis exists in the coronary artery lumen.

Exclusion criteria: patients suffering from other heart diseases, such as rheumatic heart disease, valvular heart disease or congenital heart disease; ② the following diseases, such as connective tissue disease, acute and chronic renal insufficiency, malignant tumor, or other immune inflammation participation or related diseases; ③ combined with embolic diseases, such as disseminated intravascular coagulation, pulmonary embolism, lower limb arteriovenous embolism, mesenteric arteriovenous embolism and other vascular embolic diseases; (iv) patients taking anti-inflammatory drugs, such as NSAIDS and/or steroids.

1. Experimental procedure

1.1 extraction of plasma exosomes

10mL of fasting venous blood of the patient is collected and separated to obtain serum.

Exosome isolation Step (SBI):

1) taking a 250 mu L serum sample, and melting at room temperature;

2) centrifuging at 3000g for 15min, and transferring the supernatant to a new 1.5mL Eppendorf centrifuge tube;

3) adding 67 μ L of ExoQuick, flicking the tube bottom, mixing well, standing at 4 deg.C for 30 min;

4) centrifuge at 4 ℃ for 10min at 3000g, carefully remove the supernatant, taking care not to touch the bottom pellet;

5)200 μ L Buffer B resuspend the pellet;

6) adding 200 mu L of Buffer A into the obtained heavy suspension to obtain a mixed solution;

7) taking out the purification column, cutting off the cap at the bottom (not discarding), putting the purification column into a collection tube, and centrifuging for 30 seconds at 1000 g;

8) discarding the liquid in the collecting pipe;

9) adding 500 mu L of Buffer B into the purification column, centrifuging for 30 seconds at 1000g, and discarding the liquid in the collection tube;

10) repeating the step 8) once;

11) buckling the cap cut in the step 6) at the bottom of the purification column, and adding 100 mu L of Buffer B into the purification column;

12) adding the mixed solution obtained in the step 5) into a purification column, and fully and uniformly mixing (the time can not exceed 5 min);

13) taking down the cap at the bottom of the purification column, and immediately putting the cap into a new 1.5mL Eppendorf centrifuge tube;

14) centrifuging for 30 seconds at 1000g, and obtaining the liquid in the centrifuge tube as the separated exosome.

1.2 exosome particle size analysis

Adding 1ml LPBS into the extracted exosome, and mixing uniformly. The diameter of the exosome particles was measured using a nanoparticle tracking analyzer ZetaView-particle metric.

1.3 exosome projection electron microscopy analysis

And adding 50-100 mu L of 2% paraformaldehyde solution into the extracted exosomes. And (3) dripping 5-10 mu L of exosome solution on a Formvar-carbon sample-carrying copper net, standing at room temperature for 10min, and then washing with PBS (phosphate buffer solution). Treating with 50 μ L of 1% glutaraldehyde solution on copper net for 5min, and washing with 100 μ L redistilled water for 8 times (2 min/time). Then treated with 50. mu.L of uranyl oxalate solution (pH7.0) for 5 min. Finally, treatment with 50. mu.L of methylcellulose solution was performed for 10min on ice. And (5) drying in air for 5-10 min. The copper mesh was placed in the sample box and the appropriate focus and brightness was adjusted at 80kV and photographed.

1.4 extraction of exosome RNAs

Exosome RNA was extracted using an RNA extraction kit (Qiagen, cat # 217184). Adding 600 μ L lysate into the obtained exosome stock solution, incubating at room temperature for 2min, centrifuging at 3000g for 5min, and collecting supernatant. Adding chloroform, layering, collecting upper water phase, adding 1.5 times volume of 100% ethanol, and mixing with pipette. Then, the mixture was added in portions to a column provided in the kit, and 8000g of 15s was centrifuged, and the waste liquid was discarded to recover the column. The buffer of the kit was then added to the column and the column was washed by centrifugation. Adding 80% ethanol to the column, centrifuging for 8000g for 2min, transferring the column to a new RNA-free enzyme EP tube, air drying the filter membrane on the column, adding 14 μ L of eluent, centrifuging for 8000g for 1min, discarding the column, collecting filtrate, i.e. exosome RNA, and storing in a refrigerator at-80 deg.C.

1.9 high throughput sequencing of exosome RNA

Firstly, the integrity, purity, quality and the like of the extracted exosome RNA are identified. After qualified, the cDNA first chain is synthesized by reverse transcription, then the RNA chain is degraded by RNase H, and the cDNA second chain is synthesized by taking dNTP as a raw material under the action of DNA polymerase. And (3) carrying out terminal repair, PCR amplification, purification and other steps on the purified double-stranded cDNA to obtain a sequencing library, and sequencing by using an IlluminaHiseq platform. Clean data was obtained by removing the linker, the read length containing ploy-N, and the low quality read length from the sequencing raw data. Differential expression analysis was performed on mRNA using DESeq2R package (1.16.1), Gene Ontology (GO) analysis and kyotoencyclopedia of genes and genomes (KEGG) signal pathway enrichment analysis were performed on differentially expressed transcripts by clusterierprofilerr package.

1.10 differential expression mRNA analysis

Differentially expressed mrnas were analyzed with audios software and GO and KEGG signaling pathways were analyzed with Kobas 2.0 software.

1.11 qRT-PCR detection of exosome RNA expression at mRNA level

The RNA to be detected is reversely transcribed into cDNA by utilizing a TransScript miRNA First-Strand cDNA Synthesis SuperMIX kit. Reverse transcription system: total RNA, 0.5. mu.g; 2 × TS miRNA Reaction Mix, 5 μ l; TransScript miRNA RT Enzyme Mix, 0.5 μ l; nucleic-free H₂O was added to 10. mu.l. Reaction procedure: 60min at 37 ℃ and 5s at 85 ℃. After the reverse transcription is finished, 90 μ l of nucleic-free H is added₂O is stored in a refrigerator at-20 ℃ for later use.

Designing a primer: shanghai Ouyi biomedical science and technology, Inc. and synthesized by Beijing Ongkou New Biotechnology, Inc. The primer sequences are shown in Table 1.

TABLE 1 primer sequences

Fluorescent quantitative PCR: using PerfectStartTM Green qPCR SuperMix kit in

Reactions were performed on a model 480 II fluorescent quantitative PCR instrument (Roche, Swiss). The method comprises the following steps: 2 × PerfectStartTM Green qPCR SuperMix, 5 μ l; 10 μ M Universal primer, 0.2 μ l; 10. mu.M microRNA-specific primer, 0.2. mu.l; cDNA, 1. mu.l; nuclean-free H2O, 3.6. mu.l. PCR procedure: 30s at 94 ℃; 94 ℃ for 5s, 60 ℃ for 30s, 45 cycles. Detecting product specificity by using a melting curve after circulation is finished: the temperature was slowly raised from 60 ℃ to 97 ℃ and fluorescence signals were collected 5 times per ℃ C.

The expression level was calculated by the 2-. DELTA.Ct method.

1.12 statistical methods

Statistical analysis was performed using SPSS software, with normal distribution data represented by x + -s and abnormal distribution data represented by median sum (P)₂₅,P₇₅) And (4) showing. The two groups of normal distribution data are compared by adopting a t test, the two groups of non-normal distribution data are compared by adopting a non-parameter test, and the two groups of data rates are compared by adopting a chi-square test. With P<A difference of 0.05 is statistically significant.

2. Analysis of results

2.1 venous blood was obtained from 3 ST elevation myocardial infarction (STEMI) patients (provided by the subsidiary hospital of xu zhou medical university) as STEMI group; venous blood of 3 healthy persons was used as a control group, and differential analysis of miRNA expression in peripheral blood exosomes of human subjects of STEMI group and healthy control group was performed by RNA-seq technique.

As shown in A in figure 1, the appearance and size of the patient serum exosome are observed by a transmission electron microscope, and the patient serum exosome is a circular or elliptical membrane vesicle with the diameter of 30-250nm and has a double-layer membrane structure.

The concentration and size distribution of the obtained exosomes were further analyzed using Nanoparticle Tracking Analysis (NTA) technique, and as a result, as shown in B in fig. 1, the exosome concentration of STEMI patients was measured to be 7.3 × 10⁶Per ml, mean diameter 122.9 nm.

RNA-seq technology screening 3 STEMI groups and healthy control groups human peripheral plasma exosome miRNA expression differential analysis: the differences between co-expressed peripheral plasma exosome mirnas in both groups were analyzed and it was found that there were 30 mirnas with significant differences in expression (log2 fold change >1.0, P <0.05) in the STEMI group compared to the control group, 13 of which were upregulated and 17 were downregulated (fig. 2A-2B).

2.2 screening of differentially expressed miRNAs in serum exosomes from 2.1 the top 5 miRNAs (Novel-120, hsa-miR-766-5P, Novel-54, hsa-miR-657, hsa-miR-532-5P) were subjected to qRT-PCR validation (see method 1.11 for specific parameters) in 50 additional STEMI and 50 healthy people control groups (from Xuzhou medical university Hospital and Anhui provincial Hospital), respectively, and the results are shown in FIG. 3, which indicates that the expression levels of 2 miRNAs (hsa-miR-766-5P and hsa-miR-532-5P) were significantly increased (P < 0.01) in the STEMI group compared with the control group.

2.3 drawing ROC curve of hsa-miR-766-5p expression quantity according to data obtained by 50 STEMI in 2.2 and 50 control groups of healthy people, wherein the sensitivity of hsa-miR-766-5p is 90.0%, the specificity is 90.0%, and the area under the ROC curve is 0.953; and (3) drawing a ROC curve of the expression level of the hsa-miR-532-5p (figure 4B), wherein the sensitivity of the hsa-miR-532-5p is 72.0%, the specificity is 96.0%, and the area under the ROC curve is 0.897. The sensitivity of serum blood hypersensitivity troponin for diagnosing acute myocardial infarction at present in clinic is 92.0%, the specificity is 92.0%, and the area under the ROC curve is 0.954 (figure 4C). The results show that the two miRNAs of hsa-miR-766-5p and hsa-miR-532-5p are taken as markers, and have very high sensitivity and specificity for the diagnosis of acute myocardial infarction.

2.4 KEGG pathway analysis of serum exosomes differentially expressing miRNAs: KEGG pathway annotation was performed on differentially expressed miRNAs and results are shown in fig. 5, which show that differentially expressed miRNAs participate in signaling pathway processes including NF- κ B, T cell receptor, Ras, MAPK, etc.

Those skilled in the art can make various changes, modifications and equivalent arrangements, which are equivalent to the embodiments of the present invention, without departing from the spirit and scope of the present invention, and all changes, modifications and equivalents of the technical contents disclosed above can be made; meanwhile, any changes, modifications and variations of the above-described embodiments, which are equivalent to those of the technical spirit of the present invention, are within the scope of the technical solution of the present invention.

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Claims

1. Application of miR-532-5p and/or miR-766-5p from serum exosomes in preparation of acute myocardial infarction diagnosis products.

2. Use according to claim 1, further comprising one or more of the following features:

1) miR-532-5p and/or miR-766-5p from serum exosomes is used as a biomarker;

2) the acute myocardial infarction diagnosis product is used for judging acute myocardial infarction, selecting treatment scheme and/or evaluating prognosis.

3. The use of claim 1, wherein the acute myocardial infarction diagnostic product comprises an acute myocardial infarction detection reagent, and the acute myocardial infarction detection reagent is a reagent specifically recognizing miR-532-5p and/or miR-766-5 p.

4. The use according to claim 3, wherein the agent that specifically recognizes miR-532-5p and/or miR-766-5p is selected from any one or more of the following:

(1) a primer pair for specifically amplifying miR-532-5p and/or miR-766-5 p;

(2) and the probe specifically recognizes the miR-532-5p and/or miR-766-5 p.

5. The use of claim 4, wherein when the reagent specifically recognizing miR-532-5p and/or miR-766-5p is selected from a primer pair specifically amplifying miR-532-5p and/or miR-766-5p, the primer pair specifically amplifying miR-532-5p and/or miR-766-5p comprises:

1) as shown in SEQ ID NO: 1, specifically amplifying an upstream primer sequence of miR-532-5 p;

2) as shown in SEQ ID NO: 2, a downstream primer sequence of the specific amplification miR-532-5 p;

3) as shown in SEQ ID NO: 3, and specifically amplifying an upstream primer sequence of miR-766-5 p;

4) as shown in SEQ ID NO: 4, and specifically amplifying a downstream primer sequence of the miR-766-5 p.

6. Application of a reagent for specifically recognizing miR-532-5p and/or miR-766-5p in preparation of an acute myocardial infarction diagnosis kit.

7. The use of claim 6, wherein the agent that specifically recognizes miR-532-5p and/or miR-766-5p is selected from any one or more of the following:

(1) a primer pair for specifically amplifying miR-532-5p and/or miR-766-5 p;

(2) and the probe specifically recognizes the miR-532-5p and/or miR-766-5 p.

8. The use according to claim 7, wherein, when the reagent specifically recognizing miR-532-5p and/or miR-766-5p is selected from a primer pair specifically amplifying miR-532-5p and/or miR-766-5p, the primer pair specifically amplifying miR-532-5p and/or miR-766-5p comprises:

9. An acute myocardial infarction detection kit is characterized by at least comprising an acute myocardial infarction detection reagent, wherein the acute myocardial infarction detection reagent is selected from a reagent for specifically recognizing miR-532-5p and/or miR-766-5 p.

10. The acute myocardial infarction detection kit of claim 9, wherein the reagent specifically recognizing miR-532-5p and/or miR-766-5p is selected from any one or more of the following:

(1) a primer pair for specifically amplifying miR-532-5p and/or miR-766-5 p;

(2) and the probe specifically recognizes the miR-532-5p and/or miR-766-5 p.

11. The acute myocardial infarction detection kit of claim 10, wherein when the reagent for specifically recognizing miR-532-5p and/or miR-766-5p is selected from the primer pair for specifically amplifying miR-532-5p and/or miR-766-5p, the primer pair for specifically amplifying miR-532-5p and/or miR-766-5p comprises: