CN107858355B - miRNA marker of non-alcoholic fatty liver disease and application thereof - Google Patents
miRNA marker of non-alcoholic fatty liver disease and application thereof Download PDFInfo
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Abstract
The invention provides a miRNA marker of non-alcoholic fatty liver disease and application thereof, which can be used for judging the non-alcoholic fatty liver disease. The miRNA marker is selected from: miRNA-1911 primary miRNA, miRNA-1911 precursor miRNA, and mature miRNA-1911; the miRNA-1911 primary miRNA is cleaved and expressed as mature miRNA-1911 within a human cell; the miRNA-1911 precursor miRNA is cleaved and expressed in human cells as mature miRNA-1911. The invention discovers that the miRNA marker miRNA-1911 can be used for judging the process and the severity of the non-alcoholic fatty liver disease for the first time. The test proves that the miRNA-1911 can effectively distinguish the nonalcoholic fatty liver sample from the normal sample. On the basis, the miRNA-1911 can also be used for preparing medicines for inhibiting the non-alcoholic fatty liver. The invention provides a new method for developing the non-alcoholic fatty liver disease at the molecular level in clinic, and provides a new drug target for the treatment of the non-alcoholic fatty liver disease.
Description
Technical Field
The invention belongs to the technical field of medical examination, and particularly relates to a miRNA marker of non-alcoholic fatty liver disease and application thereof.
Background
miRNA are 21-22nt non-coding RNA molecules naturally existing in vivo, and are RNA for regulating target gene expression through post-transcriptional gene silencing. It is estimated that about 1/3 genes in organisms are regulated by mirnas. The miRNA and RISC complex can be combined with a complementary sequence in a target gene mRNA5 '-UTR or 3' -UTR through base pairing to inhibit protein translation or trigger mRNA degradation, thereby negatively regulating the expression of the target gene.
Non-alcoholic fatty liver disease (NAFLD) refers to a clinical pathological syndrome characterized mainly by excessive fat deposition in liver cells due to exclusion of alcohol and other definite liver damage factors, and acquired metabolic stress liver injury closely related to insulin resistance and genetic susceptibility. Including simple fatty liver, non-alcoholic steatohepatitis and its associated cirrhosis. With the global epidemic trend of obesity and related metabolic syndrome, the nonalcoholic fatty liver disease now becomes an important cause of chronic liver disease in developed countries such as Europe and America and affluent areas of China, the prevalence rate of NAFLD of common adults is 10% -30%, wherein 10% -20% of NAFLD is nonalcoholic steatohepatitis, and the incidence rate of cirrhosis in 10 years of the NAFLD reaches 25%.
Non-alcoholic fatty liver disease can directly cause decompensated liver cirrhosis, hepatocellular carcinoma and relapse of transplanted liver, can affect the progress of other chronic liver diseases, and is involved in the onset of type 2 diabetes and atherosclerosis. Malignant tumors related to metabolic syndrome, arteriosclerotic cardiovascular and cerebrovascular diseases and liver cirrhosis are important factors influencing the quality of life and the life expectancy of non-alcoholic fatty liver patients. For this reason, non-alcoholic fatty liver disease is a new challenge in the contemporary medical field, and the harm of non-alcoholic fatty liver disease to human health will continue to increase in the near future.
miRNA is endogenous non-coding small molecular RNA for regulating gene expression, regulates gene expression at the level after transcription, and participates in physiological processes such as cell cycle, apoptosis, development, differentiation, metabolism and the like. The misexpression of miRNA in cells can cause the occurrence of various diseases including nonalcoholic fatty liver, and recent research shows that some miRNA are abnormally expressed in the blood of nonalcoholic fatty liver patients, but no consensus is achieved on which miRNA is related to the occurrence and development of nonalcoholic fatty liver. Therefore, it is necessary to find miRNA associated with the occurrence and development of non-alcoholic fatty liver disease, so as to provide an effective means for clinical diagnosis and treatment of non-alcoholic fatty liver disease.
Disclosure of Invention
In view of the above problems in the prior art, the main object of the present invention is to provide a miRNA marker for non-alcoholic fatty liver disease and application thereof, which can be used for determining non-alcoholic fatty liver disease.
In order to achieve the purpose, the invention adopts the following technical scheme: a miRNA marker of non-alcoholic fatty liver disease, selected from: miRNA-1911 primary miRNA, miRNA-1911 precursor miRNA, and mature miRNA-1911; the miRNA-1911 primary miRNA is cleaved and expressed as mature miRNA-1911 within a human cell; the miRNA-1911 precursor miRNA is cleaved and expressed in human cells as mature miRNA-1911.
As a further preference, the miRNA marker further comprises: a sequence obtained by performing base modification on miRNA-1911 or adding bases at both ends.
As a further preference, the mature miRNA-1911 comprises miRNA-1911-5p and miRNA-1911-3 p.
The invention also provides application of the miRNA marker in preparation of a tool, wherein the tool is used for predicting the risk of the non-alcoholic fatty liver and diagnosing the non-alcoholic fatty liver.
As a further preference, the means are selected from the group consisting of chips and kits; the chip comprises a solid phase carrier and an oligonucleotide probe fixed on the solid phase carrier, wherein the oligonucleotide probe comprises a part or all of a sequence specifically corresponding to miRNA-1911; the kit comprises reagents for detecting the expression level of the miRNA-1911 in a blood sample of a subject, wherein the reagents comprise primers or probes for the miRNA-1911.
As a further preference, the application method comprises:
taking a blood sample of a subject as an experimental group, and detecting the miRNA-1911 level in the blood sample of the subject;
taking a normal blood sample of the non-alcoholic fatty liver as a control group, and obtaining the miRNA-1911 level in the normal blood sample of the non-alcoholic fatty liver;
and if the miRNA-1911 level in the experimental group is lower than the miRNA-1911 level in the control group, judging that the subject is at risk of or has developed the non-alcoholic fatty liver disease.
The invention also provides application of the miRNA marker in a high-throughput sequencing platform.
As a further preference, the application method comprises:
taking a blood sample of a subject as an experimental group, and obtaining the miRNA-1911 level in the blood sample of the subject through high-throughput sequencing;
taking a normal blood sample of the non-alcoholic fatty liver as a control group, and obtaining the miRNA-1911 level in the normal blood sample of the non-alcoholic fatty liver;
and if the miRNA-1911 level in the experimental group is lower than the miRNA-1911 level in the control group, judging that the subject is at risk of or has developed the non-alcoholic fatty liver disease.
The invention also provides application of the miRNA marker in preparation of a medicine for inhibiting non-alcoholic fatty liver disease.
As a further preference, the medicament comprises the miRNA-1911 agonist.
As a further preference, said miRNA-1911 agonist is an oligonucleotide of said miRNA-1911 or said miRNA-1911 mimetic.
The invention has the beneficial effects that: the invention discovers that the miRNA marker miRNA-1911 can be used for judging the process and the severity of the non-alcoholic fatty liver disease for the first time. The test proves that the miRNA-1911 can effectively distinguish the nonalcoholic fatty liver sample from the normal sample. On the basis, the miRNA-1911 can also be used for preparing a medicine for inhibiting the non-alcoholic fatty liver and a tool for predicting the risk of the non-alcoholic fatty liver and diagnosing the non-alcoholic fatty liver. The invention provides a new method for developing the non-alcoholic fatty liver disease at the molecular level in clinic, and provides a new drug target for the treatment of the non-alcoholic fatty liver disease.
Drawings
FIG. 1 shows the expression of miRNA-1911 in the serum of non-alcoholic fatty liver disease population and normal population.
FIG. 2 shows the expression of miRNA-1911 in the serum of a mouse model of nonalcoholic fatty liver disease.
FIGS. 3a-3b are staining patterns of the hepatoma HepG2 cells after addition of oleic acid.
FIG. 4 shows the expression of miRNA-1911 in the hepatic cell line HepG2 cultured with oleic acid.
Detailed Description
The invention provides the miRNA marker of the non-alcoholic fatty liver and the application thereof, finds the miRNA related to the generation and development of the non-alcoholic fatty liver, and thus provides an effective means for clinically judging and treating the non-alcoholic fatty liver.
In order to solve the above problems, the main idea of the embodiment of the present invention is:
the miRNA marker of the non-alcoholic fatty liver disease is selected from the group consisting of: miRNA-1911 primary miRNA, miRNA-1911 precursor miRNA, and mature miRNA-1911; the miRNA-1911 primary miRNA can be sheared and expressed into mature miRNA-1911 in human cells; the miRNA-1911 precursor miRNA can be cleaved and expressed in human cells as mature miRNA-1911.
It will be appreciated that the miRNA-1911 of the present embodiments include functional equivalents of constitutive nucleic acid molecules, i.e., variants, which exhibit the same function as the intact miRNA-1911 nucleic acid molecule, although they are mutated by deletion, substitution or insertion of nucleotide residues.
It is well known in the art that in order to ensure the stability of miRNA, protective bases such as TT may be added to one or both ends of miRNA, and miRNA bases may also be modified, but the function of miRNA is not affected. Therefore, it is well known to those skilled in the art that the sequence obtained by base modification of miRNA-1911 or addition of bases at both ends without affecting the function of miRNA-1911 is also included in the protection scope of the present invention.
In some specific embodiments of the invention, the miRNA-1911 is a mature miRNA-1911. The mature miRNA-1911 comprises miRNA-1911-5p and miRNA-1911-3p, and the miRNA-1911-3p share a common seed sequence.
Although mature miRNA-1911 is used in some embodiments, one skilled in the art would expect that the primary miRNA (pi-miRNA-1911), the precursor miRNA (pre-miRNA-1911) would achieve the same technical effect as the mature miRNA-1911 because of the ability of the cell to further process the primary miRNA (pi-miRNA-1911), the precursor miRNA (pre-miRNA-1911) into the mature miRNA-1911.
The miRNA-1911 nucleic acid molecules of the present invention may exist in single-stranded or double-stranded form. Mature miRNA-1911 is predominantly in single-stranded form, whereas miRNA-1911 precursors are partially self-complementary to form a double-stranded structure. The nucleic acid molecules of the invention may be in the form of RNA, DNA, PNA, LNA.
The invention also provides application of the miRNA marker in preparation of a tool, wherein the tool is used for predicting the risk of the non-alcoholic fatty liver and diagnosing the non-alcoholic fatty liver.
Experiments in the embodiment of the invention prove that the miRNA-1911 level in urine with the non-alcoholic fatty liver is obviously lower than the miRNA-1911 level in blood, serum or plasma without the non-alcoholic fatty liver. Therefore, if the miRNA-1911 level in urine of a subject is significantly reduced compared with the miRNA-1911 level in blood, serum or plasma of non-alcoholic fatty liver, the subject can be judged to have the non-alcoholic fatty liver, so that a non-alcoholic fatty liver prevention scheme is adopted or a diagnosis basis is provided for the establishment of a clinical treatment scheme.
In addition, if the level of miRNA-1911 in the blood, serum or plasma of the subject is significant compared to the level of miRNA-1911 in blood, serum or plasma of the subject that does not develop non-alcoholic fatty liver disease, the subject is indicated to develop non-alcoholic fatty liver disease.
Further, the tool for predicting the risk of the non-alcoholic fatty liver and determining whether the non-alcoholic fatty liver occurs includes, but is not limited to, a chip and a kit. The means includes reagents for the level of expression of miRNA-1911 in the test sample. The reagent may be a primer or probe for miRNA-1911.
The chip comprises a solid phase carrier; and an oligonucleotide probe immobilized on the solid support, the oligonucleotide probe comprising a portion or all of a sequence specifically corresponding to miRNA-1911. The oligonucleotide probe can also comprise an oligonucleotide probe aiming at miRNA which is reported in the prior art and can be used for judging whether the nonalcoholic fatty liver is generated or not. The condition that the non-alcoholic fatty liver is judged by jointly detecting multiple miRNA indexes by placing the detection probes of multiple miRNAs on the same chip is also included in the protection scope of the invention.
Further, the solid phase carrier, including the solid phase carrier, can adopt various materials commonly used in the field of gene chips, such as but not limited to nylon membrane, glass or silicon slice modified by active groups (such as aldehyde group, amino group, etc.), unmodified glass slice, plastic slice, etc.
The miRNA chip may be prepared by a conventional method for manufacturing a biochip known in the art, for example, if the solid support is a modified glass slide or a silicon wafer, and the 5' end of the probe contains a poly-dT string modified with an amino group, the oligonucleotide probe may be prepared as a solution, and then spotted on the modified glass slide or the silicon wafer using a spotting apparatus, arranged into a predetermined sequence or array, and then fixed by standing overnight, so as to obtain the miRNA chip of the present invention.
The kit comprises reagents for detecting the expression level of miRNA-1911 in the blood, serum or plasma of a subject. And comparing the expression level of the miRNA-1911 in the blood, the serum or the plasma without the non-alcoholic fatty liver disease, and if the expression level of the miRNA-1911 in the blood, the serum or the plasma is detected to be remarkably reduced by the kit, judging that the non-alcoholic fatty liver disease of the subject is high in risk or has already occurred.
Further, the reagent comprises a primer and/or a probe aiming at miRNA-1911. The reagent also comprises a primer and/or a probe aiming at miRNA which is reported in the prior art and can be used for judging the risk of the non-alcoholic fatty liver or judging whether the non-alcoholic fatty liver occurs. The condition that the non-alcoholic fatty liver disease is judged by jointly detecting multiple miRNA indexes by placing the detection primers and/or probes of multiple miRNAs in the same kit is also included in the protection scope of the invention.
The embodiment of the invention also provides application of the miRNA-1911 in a high-throughput sequencing platform. The expression level of miRNA-1911 in a blood, serum or plasma sample to be detected can be obtained through high-throughput sequencing, and compared with the result of the sample to be detected, the result of the sample to be detected is easy to judge whether the sample to be detected has the risk of the non-alcoholic fatty liver or whether the sample to be detected has the non-alcoholic fatty liver or not. Therefore, the application of miRNA-1911 obtained by high-throughput sequencing and associated with nonalcoholic fatty liver is also included in the protection scope of the present invention.
The miRNA-1911 of the invention can be natural or artificial, or obtained by transfecting cells with a vector capable of expressing a DNA fragment of the miRNA-1911. The vector comprises a viral vector and a eukaryotic vector.
The viral vector may be any suitable vector, including but not limited to retroviral vectors, adenoviral vectors, adeno-associated viral vectors, herpes viral (e.g., herpes simplex, vaccinia and EB virus) vectors, alphaviral vectors.
The eukaryotic expression vector may be any suitable expression vector, including but not limited to a pCMV-Myc expression vector, pcdna3.0 expression vector, pcdna3.1 expression vector, pEGFP expression vector, pEFBos expression vector, pTet expression vector, pTRE expression vector, or a vector modified based on known expression vectors, such as pBin438, pCAMBIA1301, and the like.
The DNA fragment capable of expressing miRNA-1911 can be obtained by the following steps: searching the position of miRNA-1911 on the genome and specific sequence information from an miRNA database (http:// microrna. sanger. ac. uk/sequences), determining the position of miRNA-1911 initial miRNA according to the genome sequence, designing specific primers in the upstream and downstream 800bp intervals of the position of miRNA-1911 initial miRNA, and amplifying the sequences in the middle of the primers to obtain the DNA fragment for expressing miRNA-1911.
The invention also provides application of the miRNA-1911 in preparation of a medicine for inhibiting or treating non-alcoholic fatty liver.
The experiment proves that the miRNA-1911 is related to the nonalcoholic fatty liver, and on the basis, the miRNA-1911 can be used for inhibiting the risk of occurrence or development of the nonalcoholic fatty liver by promoting the expression of the miRNA-1911.
Further, the medicament comprises a miRNA-1911 agonist. The miRNA-1911 agonist can promote the expression of the miRNA-1911 or can activate the function of the miRNA-1911. The target for inhibition by the miRNA-1911 agonist is not limited to miRNA-1911 itself, but includes both upstream and downstream of miRNA-1911, for example: a genomic sequence encoding miRNA-1911, a miRNA-1911 target gene, a protein or a gene regulating miRNA-1911.
Further, the miRNA-1911 inhibitor comprises protein, oligonucleotide and small molecule compound.
Preferably, the miRNA-1911 inhibitor is an antisense oligonucleotide to miRNA-1911 or a miRNA-1911 mimetic.
The antisense oligonucleotide can be easily designed according to the miRNA-1911 sequence, and can obviously reduce the expression of the miRNA-1911 after the antisense oligonucleotide is transferred into a human body. "antisense oligonucleotides (AS-Ons or ASO)" also referred to AS "antisense nucleotides" refers to DNA or RNA molecules or analogs thereof having a length of about 18 to 26nt (more particularly about 19 to 22 nt).
In the present embodiment, the "antisense oligonucleotide" also includes modified antisense nucleotide obtained by means of a technique based on nucleic acid lock or nucleic acid chain skeleton modification, and the modification does not substantially change the activity of the antisense oligonucleotide, and preferably, the modification can improve the stability, activity or therapeutic effect of the antisense oligonucleotide. Nucleic acid Lock (LNA) generally refers to a modification technique that connects the 2 'oxygen atom and the 4' carbon atom of ribose via a methylene bridge. The antisense medicine developed based on the modification technology of the nucleic acid chain skeleton has greatly improved solubility, nuclease degradation resistance and other aspects, and is easy to synthesize in large amount. There are various methods for modifying the backbone of an oligonucleotide, including a thio method, for example, thio-modifying a deoxynucleotide chain to a thiodeoxynucleotide chain. The method is characterized in that oxygen atoms of phosphate bonds on a DNA skeleton are replaced by sulfur atoms, and the DNA skeleton can resist degradation of nuclease. It is understood that any modification capable of maintaining most or all of the activity of the antisense oligonucleotide is encompassed by the invention.
The drug for inhibiting non-alcoholic fatty liver disease of the present invention further comprises pharmaceutically acceptable carriers, including but not limited to: diluents, buffers, suspensions, emulsions, granules, encapsulating agents, excipients, fillers, adhesives, sprays, transdermal absorbents, wetting agents, disintegrants, absorption enhancers, surfactants, colorants, flavors, or adsorptive carriers.
The medicament can be prepared into a micro-injection, a dosage form suitable for transfection, an injection, a tablet, a powder, a granule and a capsule. The medicaments in various dosage forms can be prepared according to the conventional method in the pharmaceutical field.
The drugs may be administered alone; or in combination with other drugs capable of inhibiting non-alcoholic fatty liver disease.
The medicament may be administered ex vivo: the miRNA-1911 or the expression vector of the miRNA-1911 is introduced or transfected into human body self or variant cells in vitro, and is transported back to the human body after the in vitro cell amplification.
The medicament may be administered in vivo: miRNA-1911 or miRNA-1911 expression vectors are directly introduced into the body. Such vectors may be viral or non-viral, even naked DNA or RNA.
The subject may be a human or other mammal. More specifically, the subject is an organ, tissue, cell.
The present invention is further illustrated below with reference to specific examples, which are provided only for the purpose of illustration and are not meant to limit the scope of the present invention.
The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.
Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
Example 1
This example demonstrates that miR-1911 is low expressed in serum samples from non-alcoholic fatty liver population.
Extraction of miRNA
Using the miRNA extraction kit of Tiangen, an equal volume of lysate was added to every 200. mu.l of serum, and the mixture was shaken and mixed by an oscillator for 30 seconds. After standing at room temperature for 5min, centrifuging at 12,000rpm for 10min, collecting the supernatant, adding 200 μ l chloroform, shaking vigorously for 15 s, standing at room temperature for 5min, centrifuging at 12,000rpm for 15min, and separating the sample into three layers: yellow organic phase, intermediate layer and colorless aqueous phase, wherein RNA is mainly in the aqueous phase, transferring the aqueous phase into a new tube, slowly adding anhydrous ethanol with the volume of 1/3 volumes of the transfer solution, mixing, transferring into an adsorption column, standing at room temperature for 2min, centrifuging at 12,000rpm for 30s, and retaining the effluent. Slowly adding anhydrous ethanol with volume of 2/3 of the effluent, mixing, transferring into adsorption column, standing at room temperature for 2min, centrifuging at 12,000rpm for 30s, and retaining the adsorption column after centrifugation. Mu.l of deproteinized solution was added to the adsorption column, and centrifuged at 12,000rpm at room temperature for 30sec, and the waste liquid was discarded. 500 μ l of the rinse was centrifuged at 12,000rpm for 30 seconds at room temperature. The adsorption column was placed in a 2ml collection tube and centrifuged at 12,000rpm for 1min at room temperature to remove residual liquid. The column was then transferred to a new 1.5ml centrifuge tube, 15-30. mu.l RNase-free water was added and centrifuged at 12,000rpm for 2min at room temperature.
2. Reverse transcription
10 pg-1. mu.g of RNA template was mixed with 2. mu.l 10-fold buffer, 2. mu.l dATP (10mM), 0.5. mu.l primer, 0.5. mu.l RNase inhibitor and RNase-free water in a final volume of 20. mu.l and incubated at 37 ℃ for 1 h. Then 1. mu.l of 0.5. mu.g/. mu.l specific RT primer was added to the reaction tube, and after incubation at 70 ℃ for 5min, immediately followed by incubation on ice for at least 2min, breaking the secondary structure of RNA and primer. Finally, 20. mu.l of the above reaction mixture was mixed with 4. mu.l of 5-fold buffer, 1. mu.l of dNTP (10mM), 0.5. mu. l M-MLV reverse transcriptase, 0.5. mu.l of RNase inhibitor, 10. mu.l of polyA reaction mixture and 4. mu.l of RNase-free water, and incubated at 42 ℃ for 1 hour.
Q-PCR detection
A25. mu.l reaction system was used, with 3 parallel channels per sample, and all amplification reactions were repeated three more times to ensure the reliability of the results. The following reaction system was prepared: SYBR Green polymerase chain reaction system 12.5. mu.l, forward primer (5. mu.M/l) 1. mu.l, reverse primer (5. mu.M/l) 1. mu.l, template cDNA 2. mu.l, 8.5. mu.l enzyme-free water. All operations were performed on ice. The amplification procedure was: 95 ℃ for 10min, (95 ℃ for 20s, 60 ℃ for 55s)40 cycles. SYBR Green is used as a fluorescent marker, and PCR reaction is carried out on a fluorescent real-time quantitative PCR instrument. The band of interest was determined by melting curve analysis and electrophoresis, and relative quantification was performed by Δ Δ CT.
4. Results
As shown in figure 1, the content of miRNA-1911 in the serum of the non-alcoholic fatty liver disease population is significantly lower than that of the normal population (p <0.05), and the miRNA-1911 can be used as a marker for detecting the non-alcoholic fatty liver disease.
Example 2
This example demonstrates that miRNA-1911 is low expressed in serum of a mouse model of nonalcoholic fatty liver.
1. Non-alcoholic fatty liver animal model
Young rats (3 weeks old) were fed with a high-fat diet containing 2% cholesterol, 10% lard, 0.3% sodium cholate, etc. daily for 8 weeks. Immediately drawing whole blood, measuring the content of Total Cholesterol (TC), Triacylglycerol (TG), Free Fatty Acid (FFA), alanine Aminotransferase (ALT), aspartate Aminotransferase (AST), Triacylglycerol (TG) and Free Fatty Acid (FFA) in the blood, and increasing TC, TG, FFA, AST and ALT in the blood serum of model rats.
miRNA extraction and reverse transcription, refer to the experimental procedure in example 1.
Q-PCR detection, Experimental procedure in reference example 1
4. Results
As shown in fig. 2, in serum of the rat model of non-alcoholic fatty liver disease, the content of miRNA-1911 was significantly lower than that of rats fed with normal feed (p <0.05), and miRNA-1911 was a marker of the process of non-alcoholic fatty liver disease.
Example 3
This example demonstrates that miRNA-1911 is expressed in reduced amounts in HepG2 cells cultured with the addition of oleic acid.
1. And adding oleic acid to culture HepG2 cells, and establishing a cell model with high oil content. HepG2 cells at 2X 105And/well inoculating in six-well culture plate, and taking growth phase cell for experiment. Adding a serum-free DMEM medium, culturing for 24h, adding DMEM containing 1mM oleic acid and 10% BSA, and culturing for another 24h to establish a cell model with high oil content. The control model was cultured in serum-free DMEM medium without oleic acid and BSA. Oil red staining was used to show that the lipid content in the model cells was higher than in the control, as shown in FIGS. 3a-3 b.
miRNA extraction and reverse transcription, refer to the experimental procedure in example 1.
Q-PCR detection, Experimental procedure in reference example 1
4. Results
As shown in fig. 4, the content of miRNA-1911 was significantly lower than control cells in the oleic acid-added HepG2 cell model (p < 0.05).
The technical scheme in the embodiment of the application at least has the following technical effects or advantages:
the embodiment of the invention discovers that the miRNA marker miRNA-1911 can be used for judging the process and the severity of the non-alcoholic fatty liver disease for the first time. The test proves that the miRNA-1911 can effectively distinguish the nonalcoholic fatty liver sample from the normal sample. On the basis, the miRNA-1911 can also be used for preparing medicines for inhibiting the non-alcoholic fatty liver and prognosis and diagnosis tools. The invention provides a new method for developing the non-alcoholic fatty liver disease at the molecular level in clinic, and provides a new drug target for the treatment of the non-alcoholic fatty liver disease.
Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.
Sequence listing
<110> first Steel Hospital of Beijing university
<120> miRNA marker of non-alcoholic fatty liver disease and application thereof
<130> 2017
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 23
<212> RNA
<213> Artificial sequence ()
<400> 1
ugaguaccgc caugucuguu ggg 23
Claims (3)
1. Method for treating non-alcoholic fatty liver disease
Use of a marker in a manufacturing tool, characterized in that:
the non-alcoholic fatty liver disease
The marker is selected from:
initial
、
Precursor body
And maturation of
(ii) a The above-mentioned
Initial
Is cleaved and expressed as mature in human cells
(ii) a The above-mentioned
Precursor body
Is cleaved and expressed as mature in human cells
；
The tool is used for predicting the risk of the non-alcoholic fatty liver and diagnosing the non-alcoholic fatty liver.
2. The method of claim 1
Use of a marker in a manufacturing tool, characterized in that: the tool is selected from a chip and a kit; the chip comprises a solid phase carrier and oligonucleotide probes fixed on the solid phase carrier, wherein the oligonucleotide probes comprise specific probes which correspond to the solid phase carrier
A partial or complete sequence of (a); the reagent kit packageIncluding for detecting in a blood sample of a subject
The agent comprising a peptide directed against the expression level of
The primer or probe of (1).
3. The method of claim 1
Use of a marker in a manufacturing tool, characterized in that: the application method comprises the following steps:
using the blood sample of the subject as an experimental group, and detecting the blood sample of the subject
Horizontal;
using normal blood sample of non-alcoholic fatty liver as control group to obtain the blood sample of non-alcoholic fatty liver
Horizontal;
if in the experimental group
The level is lower than that in the control group
And (4) judging that the risk of the non-alcoholic fatty liver disease exists or the non-alcoholic fatty liver disease has already occurred.
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| CN110734971A (en) * | 2019-11-27 | 2020-01-31 | 广州中医药大学(广州中医药研究院) | group biomarker for diagnosing metabolic fatty liver mediated vascular diseases and application thereof |
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Non-Patent Citations (2)
| Title |
|---|
| Homo sapiens microRNA 1911;Bar M et al;《NCBI Reference Sequence：NR_031732.1》;20161008;origin * |
| Role of MicroRNAs in NAFLD/NASH;Gyonygi Szabo et al;《Dig Dis Sci》;20161231;摘要 * |
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