KR20200061902A - Biomarker microRNAs for diagnosing diabetic nephropathy and use thereof - Google Patents

Abstract

The present invention relates to a biomarker for diagnosing diabetic nephropathy and a use thereof and, more particularly, to a marker composition for diagnosing diabetic nephropathy comprising at least one selected from the group consisting of microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-5010), a composition and a kit for diagnosing diabetic nephropathy comprising an agent for measuring an expression level of the miRNA, and a method for diagnosing diabetic nephropathy using the marker. According to the present invention, microRNAs specific to the presence of renal complications can be useful as a biomarker for diagnosing diabetic nephropathy and predicting a progress thereof.

Description

Biomarker microRNAs for diagnosing diabetic nephropathy and use thereof

The present invention relates to a biomarker for diagnosing diabetic nephropathy and its use, more specifically, microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b) , And a marker composition for diabetic nephropathy diagnosis comprising at least one selected from the group consisting of microRNA-5010 (miR-5010), a diabetic nephropathy composition and diagnostic kit comprising an agent for measuring the expression level of the miRNA, It relates to a method for diagnosing diabetic nephropathy using the marker.

Diabetes mellitus is a type of metabolic disease, such as lack of insulin secretion or normal functioning, and is characterized by high blood sugar, which increases the concentration of glucose in the blood, and causes various symptoms and signs due to high blood sugar, and discharges glucose from urine. . Type 1 diabetes, also called pediatric diabetes, is caused by an autoimmune mechanism in which beta cells in the pancreas are destroyed, resulting in absolute or relative production deficiency of insulin, while type 2 diabetes is induced by insulin resistance and is obese, It is known to be caused by various factors such as decreased insulin receptors, decreased insulin secretion, and genetic factors.

According to the 2016 presentation of the Korean Diabetes Association, the prevalence of diabetes among adults aged 30 years or older in Korea reached a record high of 13.7% as of 2014, and the risk of concomitant disease of diabetes is also serious. 48.6% and 58.9%, and hypertension and hypercholesterolemia were also accompanied by 54.7% and 31.6%, respectively. Diabetes can cause serious complications such as cardiovascular disease, retinopathy, renal failure, and peripheral neuropathy, as well as its own risk.

Diabetic nephropathy, one of complications caused by diabetes, is a disease in which kidney function of the kidneys is damaged by high blood sugar and the like, and kidney function decreases. Diabetes progresses kidney disease slowly over 10-15 years. In the early stages of diabetes, if poorly treated, kidney function may be impaired, and in severe cases, end-stage renal failure requiring dialysis and transplantation may result. The etiology of diabetic nephropathy has not yet been clarified, but complex factors such as intrarenal hemodynamic alterations, glomerular glycolysis (glycosylation of glomerular proteins) and accumulation of sorbitol are associated with the pathogenesis of diabetic nephropathy. It is believed to be.

Currently, a renin-angiotensin system (RAS) inhibitor is mainly used to prevent diabetic nephropathy. RAS inhibitors have excellent effects in protecting kidney function by inhibiting the production of cytokines, chemokines, growth factors, etc., which induce chronic inflammatory reactions or fibrosis, as well as hemodynamic effects. However, because the genetic tendency of each patient is different, and there are side effects and resistance to treatment problems caused by long-term treatment, the therapeutic effect is less than expected. In addition, despite intensive research on the cause, diagnosis, and treatment methods for nephropathy in the past few years, a definitive treatment that can effectively treat nephrosis has not been developed. Therefore, diabetic nephropathy can be said to be the most effective way to find and treat it at an early stage.

Conventional biomarkers for diagnosing diabetic nephropathy are limited. Diagnosis of diabetic nephropathy in clinical practice is carried out regularly and repeatedly through clinical symptoms and patient history. For example, patients with long-term diabetes have nephrotic-range proteinuria and retinopathy. It is diagnosed with diabetic nephropathy if there is no other clear cause of kidney disease.

Microproteinuria (30-300 mg/d urine albumin for 24 hours or albumin excretion rate 20-200 ug/min) is most frequently used as a biomarker to predict the progression of diabetic nephropathy early and to predict vascular complications other than nephropathy. . According to existing studies, microproteinuria is a risk factor predicting a decrease in glomerular filtration rate and an independent risk factor for progressing to chronic renal failure, so it is currently recommended to measure microproteinuria in diabetes diagnosis and treatment guidelines. However, recent studies have shown that the frequency of natural loss of microproteinuria in diabetic patients is high (Diabetes Care. 2001 Sep;24(9):1560-6.), and the renal function is reduced without microproteinuria (glomerular filtration rate <60 ml/min/ 1.73m ² ) Due to the high frequency of diabetic patients, it has been reported that there is a limitation in diagnosing diabetic nephropathy only with microproteinuria and judging it as a progression indicator.

Accordingly, the present inventors analyzed miRNAs present in the serum-derived exosomes of the subject in order to discover a biomarker capable of effectively diagnosing diabetic nephropathy, and significantly increased expression in the diabetic patient group with renal complications compared to the normal control group. The present invention was completed by discovering four types of miRNAs.

In order to solve the above-described conventional problems, the present inventors collect serum from subjects corresponding to the normal group, the diabetic group without nephropathy, and the diabetic group with nephropathy and extract exosome RNA therefrom. As a result of comparing and analyzing the expression level of the exosome microRNA through NGS analysis after separation, 4 types of microRNAs in which the expression level was significantly increased only in the diabetic group with renal complications compared to the normal group were found. The present invention was completed.

Accordingly, the present invention is selected from the group consisting of microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-5010). It is an object of the present invention to provide a marker composition for diabetic nephropathy diagnosis.

In addition, the present invention is selected from the group consisting of microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-5010). Another object is to provide a composition for diagnosing diabetic nephropathy and a kit for diagnosing diabetic nephrosis comprising the composition, comprising an agent that measures one or more expression levels.

In addition, the present invention provides microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-) in biological samples derived from a subject. Another object is to provide an information providing method for diagnosing diabetic nephropathy, comprising the step of measuring one or more expression levels selected from the group consisting of 5010).

However, the technical problem to be achieved by the present invention is not limited to the above-mentioned problems, and other problems that are not mentioned will be clearly understood by those skilled in the art from the following description.

In order to achieve the object of the present invention as described above, the present invention is microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 It provides a marker composition for diabetic nephropathy diagnosis, comprising at least one selected from the group consisting of (miR-5010).

In addition, the present invention is selected from the group consisting of microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-5010). It provides a composition for diagnosing diabetic nephropathy, comprising an agent for measuring one or more expression levels.

In addition, the present invention provides a diabetic nephropathy diagnostic kit comprising the composition for diagnosing diabetic nephropathy.

In one embodiment of the present invention, the miR-642a may be composed of the nucleotide sequence of SEQ ID NO: 1.

In another embodiment of the present invention, the miR-1246 may be composed of the nucleotide sequence of SEQ ID NO: 2.

In another embodiment of the present invention, the miR-1255b may be composed of the nucleotide sequence of SEQ ID NO: 3.

In another embodiment of the present invention, the miR-5010 may be composed of the nucleotide sequence of SEQ ID NO: 4.

In another embodiment of the present invention, the agent for measuring the expression level may be a sense and antisense primer, or a probe that complementarily binds to the microRNA.

In addition, the present invention provides microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-) in biological samples derived from a subject. It provides a method for providing information for diagnosing diabetic nephropathy, comprising the step of measuring one or more expression levels selected from the group consisting of 5010).

In one embodiment of the present invention, the biological sample may be blood or serum-derived exosomes.

In another embodiment of the present invention, the expression level is next generation sequencing (NGS), polymerase chain reaction (PCR), reverse transcriptase chain reaction (RT-PCR), real-time polymerase chain reaction (Real) -time PCR), RNase protection assay (RNase protection assay; RPA), microarray (microarray), and may be measured through one or more methods selected from the group consisting of Northern blotting (northern blotting).

The present inventors analyzed the microRNA contained in the exosomes isolated from the blood of the normal group, the diabetic group without nephropathy, and the diabetic group with nephropathy, and the expression was significant only in the diabetic group with nephropathy compared to the normal group. The specific microRNAs miR-642a, miR-1246, miR-1255b, and miR-5010 were identified for the presence or absence of renal complications. Accordingly, the discovered microRNAs may be useful as a biomarker for diagnosing diabetic nephropathy and predicting progress.

1 is a result of comparative analysis of the expression level of microRNAs in exosomes isolated from the serum of normal group (HV) and diabetic group without nephropathy (Nephro-X), and FIG. 1a shows renal complications compared to normal group. Heat map results of hierarchical clustering according to the expression level of microRNAs whose expression was significantly reduced or increased by more than 2 times (p <0.05) in the diabetic group without, and FIG. 1B shows the expression level of the detected microRNAs. It is a scatter plot result.
2 is a result of comparative analysis of the expression level of microRNAs in exosomes isolated from serum of normal (HV) and diabetic (Nephro-O) subjects with nephropathy, and FIG. 2A shows nephropathy compared to the normal group. In the diabetic group, a heat map result of hierarchical clustering according to the expression level of microRNAs whose expression is significantly reduced or increased by 2 times or more (p<0.05) is shown, and FIG. 2b is according to the expression level of the detected microRNAs. It is a scatter plot result.
Figure 3 is a result of comparative analysis of the expression level of microRNAs in exosomes isolated from the serum of subjects with nephropathy without diabetes (Nephro-X) and nephropathy (Nephro-O), Figure 3a Heat map results of hierarchical clustering according to the expression level of microRNAs whose expression was significantly reduced (p <0.05) more than 2 times (p <0.05) in diabetic group with renal complications compared to diabetic group without complications, and FIG. It is a scatter plot result according to the expression level of the detected microRNAs.
4 is a multi-dimensional scale (MDS) for the normal group (HV), the diabetic group without nephropathy (Nephro-X) and the diabetic group with nephropathy (Nephro-O), based on the results of FIGS. 1 to 3. It is the result of the analysis.
5 is a result of deriving only microRNAs with significantly increased expression only in the diabetic group with renal complications compared to the normal group by expressing only the microRNAs with significantly increased expression from the results of FIGS.
Figure 6 hsa-miR-642a-3p, hsa-miR-1246, hsa-miR-1255b-5p and hsa-miR-5010-5p between normal group (HV) and diabetic group with nephropathy (Nephro-O) It is a result of measuring and comparing the expression level of.

The present inventors have conducted diligent research to discover biomarkers for diagnosing diabetes. As a biomarker that can be used to diagnose diabetic nephropathy, microRNA-642a (miR-642a), microRNA-1246 (miR-1246), and micro The present invention was completed by discovering RNA-1255b (miR-1255b) and microRNA-5010 (miR-5010).

Accordingly, the present invention is selected from the group consisting of microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-5010). It provides a marker composition for diabetic nephropathy diagnosis, comprising at least one.

In addition, the present invention is selected from the group consisting of microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-5010). Provided is a diabetic nephropathy diagnosis composition comprising the agent for measuring at least one expression level, and a diabetic nephropathy diagnosis kit comprising the diagnostic composition.

Thus, the miR-642a, more preferably hsa-miR-642a-3p, which is a biomarker for diagnosing diabetic nephropathy discovered in the present invention, may be composed of a nucleotide sequence represented by SEQ ID NO: 1 (NCBI, MIMAT0020924). At this time, it may include a base sequence having a sequence homology of 70% or more, preferably 80% or more, more preferably 90% or more, and most preferably 95% or more with the base sequence represented by SEQ ID NO: 1 .

In addition, the miR-1246, more preferably hsa-miR-1246, which is a biomarker for diagnosing diabetic nephropathy discovered in the present invention, may be composed of a nucleotide sequence represented by SEQ ID NO: 2 (NCBI, MIMAT0005898). At this time, it may include a base sequence having a sequence homology of 70% or more, preferably 80% or more, more preferably 90% or more, and most preferably 95% or more with the base sequence represented by the SEQ ID NO: 2 .

In addition, the miR-1255b, more preferably hsa-miR-1255b-5p, which is a biomarker for diagnosing diabetic nephropathy discovered in the present invention, may be composed of a nucleotide sequence represented by SEQ ID NO: 3 (NCBI, MIMAT0005945). At this time, it may include a base sequence having a sequence homology of 70% or more, preferably 80% or more, more preferably 90% or more, and most preferably 95% or more with the base sequence represented by the SEQ ID NO: 3 .

In addition, the biomarker for diagnosing diabetic nephropathy discovered in the present invention, the miR-5010, more preferably hsa-miR-5010-5p may be made of a nucleotide sequence represented by SEQ ID NO: 4 (NCBI, MIMAT0021043). At this time, it may include a base sequence having a sequence homology of 70% or more, preferably 80% or more, more preferably 90% or more, and most preferably 95% or more with the nucleotide sequence represented by SEQ ID NO:4. .

As used in the present invention, the term “diagnosis” means broadly determining the condition of a patient's illness across all aspects. The content of the judgment is the name of the disease, the etiology, the type of illness, the severity, the detailed aspects of the bed, and the presence or absence of complications. Diagnosis in the present invention is to determine whether the onset of diabetes complications and progression level, and more preferably, the diabetes complications may be diabetic nephropathy.

In the present invention, the formulation measuring the expression level of the miR-642a, miR-1246, miR-1255b and/or miR-5010 may be sense and antisense primers or probes that complementarily bind to each of the microRNAs. However, it is not limited thereto.

The term “primer” used in the present invention is a short gene sequence serving as a starting point for DNA synthesis, and means an oligonucleotide synthesized for the purpose of diagnosis, DNA sequencing, and the like. The primers can be generally synthesized to a length of 15 to 30 base pairs, but may vary depending on the purpose of use, and can be modified by methylation, capping, or the like by a known method.

As used in the present invention, the term “probe” refers to a nucleic acid capable of specifically binding mRNA of several bases to hundreds of bases produced through enzymatic chemical separation or synthesis. The presence or absence of mRNA can be confirmed by labeling radioactive isotopes or enzymes, and can be designed and modified in a known manner.

The diagnostic kit of the present invention consists of one or more other component compositions, solutions or devices suitable for analytical methods.

For example, in order to perform PCR, the kit of the present invention contains genomic DNA derived from a sample to be analyzed, a primer set specific for the marker gene of the present invention, an appropriate amount of DNA polymerase, dNTP mixture, PCR buffer solution and water. It may be a kit including. The PCR buffer solution may contain KCl, Tris-HCl and MgCl ₂ . In addition, the components necessary for performing electrophoresis capable of confirming whether or not the PCR product is amplified may be additionally included in the kit of the present invention.

In addition, the kit of the present invention may be a kit including essential elements necessary for performing RT-PCR. RT-PCR kits include test tubes or other suitable containers, reaction buffers, enzymes such as deoxynucleotides (dNTPs), Taq-polymerases and reverse transcriptases, DNase, RNase inhibitors, DEPCs, in addition to each primer pair specific for a marker gene. -It may include DEPC-water, sterile water, and the like. In addition, it may include a primer pair specific to the gene used as a quantitative control.

The present inventors discovered and identified the miR-642a, miR-1246, miR-1255b, and miR-5010 as biomarkers for diabetic nephropathy diagnosis through specific examples.

In one embodiment of the present invention, the subjects of the normal group, the diabetic group without nephropathy and the diabetic group with nephropathy are recruited and their serum is provided to separate the exosomes from the serum and express the miRNA in the exosomes. Levels were compared (see Example 1).

In another embodiment of the present invention, the difference between the normal group and the diabetic group without nephropathy, the diabetic group with normal and nephropathy, the diabetic group without nephropathy and the diabetic group with nephropathy, respectively, were analyzed. As a result of analyzing the expression level of the microRNAs using a multi-dimensional scale method, it was found that the normal group and the diabetic group were distinguished in two dimensions, and there was no significant difference in the presence or absence of renal complications. In addition, the microRNAs showing significant increase in expression in each of the comparative analysis results were shown and compared, and in detail, in 40 miRNAs with increased expression in the diabetic group with renal complications compared to the normal group, height compared to the normal group Eight types except for the common 32 miRNAs with increased expression in the diabetic group without complications were selected, and one of the eight miRNAs with increased expression in the diabetic group with nephropathy compared to the diabetic group without nephropathy. Four miRNAs, which showed the most significant expression difference among the seven, were finally selected. The hsa-miR-642a-3p, hsa-miR-1246, hsa-miR-1255b-5p and hsa-miR-5010-5p screened above actually showed significant differences in expression between the normal group and the diabetic group with nephropathy. It was confirmed that it was shown (Examples 2-1 to 2-4).

Accordingly, as another aspect of the present invention, the present invention provides microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and micro It provides a method for providing information for diabetic nephropathy diagnosis, comprising the step of measuring one or more expression levels selected from the group consisting of RNA-5010 (miR-5010).

The biological sample derived from the subject may include tissue, cells, whole blood, blood, saliva, sputum, cerebrospinal fluid, or urine, and more preferably blood or serum-derived exosomes, but is not limited thereto.

In the present invention, if the expression level of miR-642a, miR-1246, miR-1255b and/or miR-5010 in exosomes isolated from a subject-derived sample according to the method is higher than normal, it can be determined as diabetic nephropathy. have.

The expression level of the microRNAs is a polymerase chain reaction (PCR), reverse transcriptase chain reaction (RT-PCR), real-time polymerase chain reaction (Real-time PCR), RNase protection assay (RNase) in a conventional manner known in the art. protection assay; RPA), microarray, or Northern blotting (northern blotting) may be measured by one or more methods selected from the group consisting of, but is not limited to.

Hereinafter, preferred embodiments are provided to help understanding of the present invention. However, the following examples are only provided to more easily understand the present invention, and the contents of the present invention are not limited by the following examples.

[Example]

Example 1. Experiment preparation and experiment method

1-1. Recruitment for analysis

The present inventors, in order to discover biomarkers capable of diagnosing diabetic nephropathy due to diabetes at an early stage, each of 18 normal groups (HV), 33 diabetic patients without nephropathy (Nephro-X), and 23 patients with nephropathy (Nephro-O) were recruited, and serum samples were taken from them to find biomarkers showing differences between groups. To this end, serum was provided from subjects in each group.

1-2. RNA isolation and sequencing from serum

Exoquick (SBI, USA) was used to separate exosomes from serum samples collected from subjects of each group in Example 1-1, and total RNA was extracted from the separated exosomes using Qiagen miRNeasy kit. Was extracted, and the quality of the extracted total RNA was measured according to the manufacturer's instructions using Agilent RNA Bioanalyzer.

Thereafter, using the isolated RNA, a library was prepared using a Takara SMARTer smRNA for illumina kit from Macrogen, and then a next-generation sequencing (NGS) was performed with Hiseq 2500 to detect microRNA.

Example 2. Discovery of microRNA for diabetic nephropathy diagnosis

2-1. Normal group vs. Analysis of microRNA with difference in expression level of diabetic group without nephropathy

First, the present inventors to identify the microRNA (miRNA) having a difference in the expression level in the blood-derived exosomes of 18 normal groups and 33 diabetic groups without renal complications, the methods of Examples 1-1 and 1-2 According to the results, exosomes were separated from serum collected from each subject, and NGS analysis was performed on the extracted total RNA. As a result of the analysis, 107 miRNAs were detected, and further, the expression levels of the detected miRNAs were compared.

As a result, as shown in FIG. 1A, 62 miRNAs expressing at least twice (p<0.05) expression in the diabetic group (Nephro-X) without nephropathy compared to the normal group (HV) among the 107 miRNAs are 62 There were 45 increasing miRNAs. In addition, scatter plot results according to the expression level of miRNAs showing the significant expression change are shown in FIG. 1B.

2-2. Normal group vs. Analysis of microRNA with difference in expression level of diabetic group with nephropathy

In addition to the above results, the present inventors used the same method as above to identify miRNAs having different levels of expression in blood-derived exosomes of 18 normal groups and 23 diabetic groups with kidney complications. An NGS analysis was performed. As a result of analysis, 95 miRNAs were detected, and further, the expression levels of the detected miRNAs were compared.

As a result, as shown in FIG. 2A, 55 miRNAs expressing at least twice (p<0.05) expression in the diabetic group (Nephro-O) with nephropathy compared to the normal group (HV) among 95 miRNAs were expressed. There were 40 increasing miRNAs. In addition, scatter plot results according to the expression level of miRNAs showing the significant expression change are shown in FIG. 2B.

2-3. Diabetic group without nephropathy vs. Analysis of miRNA with difference in expression level of diabetic group with nephropathy

In addition, the present inventors in order to identify miRNAs having different levels of expression in blood-derived exosomes of 33 diabetic groups without renal complications and 23 diabetic groups with renal complications, total RNA in exosomes obtained by the same method as above. The NGS analysis was performed on. As a result of analysis, 12 miRNAs were detected, and further, the expression levels of the detected miRNAs were compared.

As a result, as shown in FIG. 3A, the expression of at least 2 times (p <0.05) decreased in the diabetic group with nephropathy (Nephro-O) compared to the diabetic group without nephropathy (Nephro-X) among 12 miRNAs. There were 9 miRNAs and 3 miRNAs with increased expression. In addition, scatter plot results according to the expression level of miRNAs showing the significant expression change are shown in FIG. 3B.

2-4. Normal group vs. Diabetic group without nephropathy vs. Analysis of miRNAs with differences in expression levels between diabetic groups with nephropathy

Furthermore, for the normal group, the diabetic group without nephropathy, and the diabetic group with nephropathy, a change amount for each sample was derived through multidimensional scaling (MDS) and visualized and analyzed. As a result, as shown in FIG. 4, the two-dimensional spatial location of each of the normal group and the diabetic group was clearly divided into two groups, but the difference according to the presence or absence of renal complications was not significantly divided. However, if a miRNA that differs depending on the presence or absence of renal disease is found, it can be used as a useful biomarker for diagnosing renal disease, and the present inventors are based on the analysis results obtained through the results of Examples 2-1 to 2-3. Therefore, additional comparative analysis was conducted to discover miRNAs for diagnosing renal complications.

To this end, miRNAs identified as having a significant increase in expression as a result of each analysis are shown in the Venn diagram of FIG. 5. As a result, 40 miRNAs (C/A) with significantly increased expression in diabetic-diabetic group (Diabetes-Nephro-O) compared to normal group (HV), and diabetic group without nephropathy compared to normal group ( Diabetes-Nephro-X) compared to the normal group by comparing 45 miRNAs (B/A) with significantly increased expression, excluding 32 miRNAs (C/A & B/A), which have renal complications compared to the normal group. Eight miRNAs with increased expression were obtained only in the diabetic group. Thereafter, in Example 2-3, compared with the obtained 8 miRNAs, 3 miRNAs (C/B) results in significantly increased expression in the diabetic group with renal complications compared to the diabetic group without renal complications, are common. As a result, one miRNA with increased expression was identified. Thus, 7 types except for the common one type of miRNAs were obtained from the eight miRNAs, and four types of miRNAs showing the most significant difference in expression levels were finally selected. The selected four types of miRNAs are shown in Table 1 below, and it was confirmed that 2.51, 2.02, 2.08, and 2.54 times the difference in renal complications compared to the normal group.

Diabetes-kidney syndrome/normal group (C/A)
fold change hsa-miR-642a-3p 2.51 * hsa-miR-1246 2.02 ** hsa-miR-1255b-5p 2.08 ** hsa-miR-5010-5p 2.54 **

In addition, as shown in Figure 6, the normal group (HV) and diabetic group with nephropathy (Nephro-O) 4 miRNAs found above, hsa-miR-642a-3p, hsa-miR-1246, It was confirmed that the expression of hsa-miR-1255b-5p and hsa-miR-5010-5p showed a significant difference.

Therefore, the above results show that four miRNAs, each of which significantly increased expression only in the diabetic group with nephropathy, can be usefully used as a biomarker for diagnosing diabetic nephropathy compared to normal people.

The above description of the present invention is for illustration only, and those skilled in the art to which the present invention pertains can understand that the present invention can be easily modified into other specific forms without changing the technical spirit or essential features of the present invention. There will be. Therefore, it should be understood that the embodiments described above are illustrative in all respects and not restrictive.

<110> Soonchunhyang University Industry Academy Cooperation Foundation <120> Biomarker microRNAs for diagnosing diabetic nephropathy and use          thereof <130> PD18-145 <160> 4 <170> KoPatentIn 3.0 <210> 1 <211> 22 <212> RNA <213> hsa-miR-642a-3p <400> 1 agacacauuu ggagagggaa cc 22 <210> 2 <211> 19 <212> RNA <213> hsa-miR-1246 <400> 2 aauggauuuu uggagcagg 19 <210> 3 <211> 22 <212> RNA <213> hsa-miR-1255b-5p <400> 3 cggaugagca aagaaagugg uu 22 <210> 4 <211> 22 <212> RNA <213> hsa-miR-5010-5p <400> 4 agggggaugg cagagcaaaa uu 22

Claims (19)

Contains one or more selected from the group consisting of microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-5010) The marker composition for diabetic nephropathy diagnosis.
According to claim 1,
The miR-642a is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 1, a marker composition.
According to claim 1,
The miR-1246 is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 2, a marker composition.
According to claim 1,
The miR-1255b is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 3, a marker composition.
According to claim 1,
The miR-5010 is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 4, a marker composition.
One or more expression levels selected from the group consisting of microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-5010) The composition for diagnosing diabetic nephropathy, comprising an agent for measuring the.
The method of claim 6,
The miR-642a is characterized by consisting of the nucleotide sequence of SEQ ID NO: 1, diabetic nephropathy diagnostic composition.
The method of claim 6,
The miR-1246 is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 2, diabetic nephropathy composition.
The method of claim 6,
The miR-1255b is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 3, diabetic nephropathy composition.
The method of claim 6,
The miR-5010 is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 4, diabetic nephropathy composition.
The method of claim 6,
The agent for measuring the expression level is characterized in that the sense and antisense primers, or probes that complementarily bind to the microRNA, diabetic nephropathy diagnostic composition.
A kit for diagnosing diabetic nephropathy comprising the composition of any one of claims 6 to 11.
Group consisting of microRNA-642a (miR-642a), microRNA-1246 (miR-1246), microRNA-1255b (miR-1255b), and microRNA-5010 (miR-5010) in biological samples from a subject Method for providing information for diagnosing diabetic nephropathy, comprising the step of measuring one or more expression levels selected from.
The method of claim 13,
The miR-642a is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 1, information providing method.
The method of claim 13,
The miR-1246 is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 2, information providing method.
The method of claim 13,
The miR-1255b is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 3, information providing method.
The method of claim 13,
The miR-5010 is characterized in that consisting of the nucleotide sequence of SEQ ID NO: 4, information providing method.
The method of claim 13,
The biological sample is blood or serum-derived exosome, characterized in that, information providing method.
The method of claim 13,
The expression level is next generation sequencing (NGS), polymerase chain reaction (PCR), reverse transcriptase chain reaction (RT-PCR), real-time polymerase chain reaction (Real-time PCR), RNase protection assay (RNase protection assay; RPA), microarray (microarray), and Northern blotting (northern blotting) characterized in that it is measured by one or more methods selected from the group consisting of, information providing method.

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