Application of miR-15b-5p in preparation of endometriosis diagnostic agent and diagnostic kit
By detecting the expression level of miR-15b-5p in serum exosomes and utilizing Q-PCR technology and an exosome capture device, the shortcomings of traditional diagnostic methods have been overcome, achieving a non-invasive, efficient, and specific diagnosis of endometriosis, thus improving the accuracy and safety of diagnosis.
Patent Information
- Application Number
- CN202511539665.4
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-10-27
- Publication Date
- 2026-01-20
AI Technical Summary
Existing technologies cannot provide a non-invasive, efficient, and specific diagnostic method for endometriosis. Traditional diagnostic methods such as ultrasound examination and serum CA125 markers have insufficient sensitivity and specificity, while laparoscopy is invasive and expensive, failing to meet the needs of routine screening.
Using miR-15b-5p expression detection reagent, combined with Q-PCR technology and an exosome capture device, the expression level of miR-15b-5p in serum exosomes was detected. After capturing exosomes using a porous affinity microfluidic exosome capture chip and a PDMS nickel foam scaffold, miRNA was detected. A relative expression threshold of 1.204 was set for diagnosis.
It achieves high sensitivity (0.908) and high specificity (0.881) in the diagnosis of endometriosis, with an AUC value of 0.898 for the ROC curve, demonstrating good diagnostic value and meeting the needs for non-invasive and efficient diagnosis.
Smart Images
Figure CN121362828A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The application belongs to the field of endometriosis diagnosis, and particularly relates to application of a miR-15b-5p expression amount detection agent in preparation of an endometriosis diagnosis agent and an endometriosis diagnosis kit. BACKGROUND
[0002] Endometriosis (endometriosis) is a common gynecological benign disease, which has the characteristics of invasion, metastasis and recurrence similar to malignant tumors, and the incidence rate is as high as 10%-15% in women of childbearing age, and shows a rising trend year by year. Endometriosis not only causes patients to have symptoms such as progressive and aggravated dysmenorrhea, chronic pelvic pain, and dyspareunia, but also seriously affects the reproductive function. According to statistics, about 30%-50% of endometriosis patients have infertility, which brings great distress to the physical and mental health and quality of life of patients, and also causes heavy social and medical burden.
[0003] At present, the diagnosis of endometriosis faces many challenges. In the traditional diagnosis method, although the ultrasonic examination is a non-invasive method, the detection rate for small lesions or deep infiltrating endometriosis is low, and misdiagnosis and missed diagnosis are easy to occur; the specificity and sensitivity of serum tumor markers such as CA125 are insufficient, and it is difficult to serve as a diagnosis basis. Laparoscopy combined with pathological biopsy is the current "gold standard" for the diagnosis of endometriosis, but this method is a invasive operation, which has problems such as anesthesia risk, surgical complications, and high cost, and cannot be used as a routine screening method, and it is also difficult to meet the needs of patients for non-invasive diagnosis.
[0004] Therefore, finding a non-invasive, efficient and specific endometriosis diagnosis method has become a key problem to be solved in the current gynecological medical field.
[0005] Exosomes, as a kind of vesicular structure with a diameter of about 30-150 nm secreted by cells, widely exist in body fluids such as blood, urine and ascites, can carry nucleic acids, proteins, lipids and other bioactive substances of the source cells, and play an important role in cell-to-cell information transmission and disease development. Because exosomes have good biological stability, and the contents can reflect the pathological and physiological state of the source cells, they are considered as a very potential disease diagnosis biomarker carrier.
[0006] MicroRNAs (miRNAs) are a class of non-coding RNAs with a length of about 20-24 nucleotides, which can participate in cell proliferation, apoptosis, invasion and other biological processes by regulating the expression of target genes. Studies have found that the expression profile of miRNA will change significantly under disease conditions, and it is stable in body fluid exosomes. In recent years, more and more studies have shown that the expression level of exosome miRNA in the body fluid (such as serum and ascites) of endometriosis patients is significantly different from that of healthy people, and some specific exosome miRNAs can not only accurately distinguish endometriosis patients from healthy controls, but also reflect the severity and stage of the disease.
[0007] At present, although relevant studies have found many potential exosome miRNA markers for endometriosis, most of the research sample sizes are small, and there is a lack of unified detection standards and diagnostic thresholds, so an endometriosis non-invasive diagnosis model based on exosome miRNA has not yet been formed for clinical promotion. SUMMARY
[0008] To solve the above problems, the application provides an application of an expression amount detection agent of miR-15b-5p in preparation of an endometriosis diagnosis agent.
[0009] In one specific embodiment, the expression amount detection agent of miR-15b-5p is a Q-PCR detection agent.
[0010] The application further provides an endometriosis diagnosis kit, comprising an expression amount detection agent of miR-15b-5p.
[0011] In one specific embodiment, the expression amount detection agent of miR-15b-5p is a Q-PCR detection agent.
[0012] In one specific embodiment, the diagnosis kit further comprises an exosome capture device.
[0013] In one specific embodiment, the expression amount detection agent of miR-15b-5p is used to detect the expression amount of miR-15b-5p in exosomes from serum.
[0014] In one specific embodiment, the diagnosis kit further comprises a negative control.
[0015] Compared with the control group, the relative expression amount threshold of miR-15b-5p is 1.204, and when the relative expression amount of miR-15b-5p is greater than 1.204, the subject is most likely to have endometriosis.
[0016] The application uses the miR-15b-5p expression amount in serum exosomes as a diagnostic marker of endometriosis, the sensitivity is 0.908, the specificity is 0.881, the false positive rate is 0.119, the false negative rate is 0.092, and the AUC value of the ROC curve is 0.898. It can be seen that the sensitivity and specificity of miR-15b-5p are both high, and it has good diagnostic value. BRIEF DESCRIPTION OF DRAWINGS
[0017] Figure 1 The miR-15b-5p expression amount in serum exosomes of endometriosis and healthy controls.
[0018] Figure 2 The ROC curve of the miR-15b-5p expression amount in serum exosomes as an endometriosis diagnostic index. DETAILED DESCRIPTION
[0019] The principles and characteristics of the application are described below in conjunction with the accompanying drawings, and the examples are only used to explain the application and are not used to limit the scope of the application.
[0020] 1. Detecting endometriosis patient exosome differential miRNA by using porous affinity microfluidic exosome capture chip
[0021] The support of the chip is made of PDMS and foamed nickel, and the support is modified by streptavidin and biotinylated exosome protein antibody to obtain a chip that can specifically capture exosomes.
[0022] The venous blood of the endometriosis patient is centrifuged at 3000 rpm / min for 10 min to obtain the patient's plasma, which is placed in a-80℃ refrigerator for use.
[0023] The patient's plasma is passed into the chip at a flow rate of 10 μl / min, and 100 μl of plasma is passed into each chip, and then PBS is washed three times. At this time, the exosomes have been captured on the support in the chip.
[0024] The TRIZOL method is used to pass into the chip, and the RNA of the exosomes in the capture chip is captured. The obtained RNA is reverse transcribed and subjected to Q-PCR to obtain the cq value (u6 as an internal reference), and the relative abundance is expressed by 2^-ΔCt / 2^-ΔΔCt. The obtained cq value is statistically analyzed to obtain the differential expression result of miRNA.
[0025] The case group and the control group are combined for a total of n=120; the plasma exosomes are enriched by the PAE-CM microfluidic chip, and then quantified by RT-qPCR.
[0026] The results are as follows Figure 1As shown, 7 miRNAs were found to be directionally stable and significantly different between cases vs controls, of which 1 miRNA was miR-15b-5p.
[0027] 2. Diagnostic value of miR-15b-5p as a single indicator
[0028] Using the relative expression amount of miR-15b-5p (endometriosis patient / healthy control threshold 1.204) to diagnose endometriosis (sample size 124, of which 65 patients and 59 healthy people), calculate the specificity, sensitivity, true positive rate, false positive rate, AUC value and other key data, and make a ROC curve.
[0029] The results show that there are 59 true positives, 52 true negatives, 7 false positives and 6 false negatives; the sensitivity is 0.908, the specificity is 0.881, the false positive rate is 0.119, the false negative rate is 0.092, and the ROC curve is as shown in Figure 2 The AUC value is 0.898. It can be seen that the sensitivity and specificity of miR-15b-5p are both high, and it has good diagnostic value.
[0030] The above only describes the preferred embodiments of the present application and is not intended to limit the present application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principles of the present application shall be included in the protection scope of the present application.
Claims
1. Use of an expression amount detection agent of miR-15b-5p in preparation of a diagnostic agent for endometriosis.
2. Use according to claim 1, characterized in that, The expression amount detection agent of miR-15b-5p is a Q-PCR detection agent.
3. A diagnostic kit for endometriosis, characterized by, The expression amount detection agent of miR-15b-5p.
4. The endometriosis diagnostic kit according to claim 3, characterized in that, The expression amount detection agent of miR-15b-5p is a Q-PCR detection agent.
5. The endometriosis diagnostic kit according to claim 4, characterized in that, Also included is an exosome capture device.
6. The endometriosis diagnostic kit according to claim 5, characterized in that, The expression amount detection agent of miR-15b-5p is used to detect the expression amount of miR-15b-5p in exosomes from serum.
7. The endometriosis diagnostic kit according to claim 6, characterized in that, Also included is a negative control.
8. The endometriosis diagnostic kit according to claim 7, characterized in that, The threshold value of the relative expression amount of miR-15b-5p is 1.2 compared with the control group, and when the relative expression amount of miR-15b-5p is greater than 1.2, the subject is very likely to have endometriosis.