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Application of hsa_circ_0003764 as marker to preparation of diagnosis preparation for intrauterine adhesion

A technology for intrauterine adhesions and diagnostic reagents, applied in biochemical equipment and methods, microbe measurement/testing, drug combinations, etc., can solve problems such as functions that have not been reported in research

Active Publication Date: 2019-06-07
NANJING DRUM TOWER HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, 4 literatures have reported the existence of circPTPN12, but its function has not yet been reported

Method used

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  • Application of hsa_circ_0003764 as marker to preparation of diagnosis preparation for intrauterine adhesion
  • Application of hsa_circ_0003764 as marker to preparation of diagnosis preparation for intrauterine adhesion
  • Application of hsa_circ_0003764 as marker to preparation of diagnosis preparation for intrauterine adhesion

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Expression of hsa_circ_0003764 in endometrial scar tissue.

[0027] 1. Materials, reagents, equipment

[0028] 1.1 Source of human endometrial tissue

[0029] 7 cases of normal endometrial tissue and 6 cases of endometrial tissue with severe intrauterine adhesion syndrome were collected. When obtaining endometrial tissue, the size of follicles was detected by ultrasound to ensure that the endometrial tissue acquisition stage was unified into the late proliferation stage. All participants signed a written informed consent form, which was approved by the Ethics Committee of Nanjing Drum Tower Hospital.

[0030] 1.2 Main reagents

[0031] Trizol (Invitrogen), Reverse Transcriptase (Takara), Qpcrase (Roche).

[0032] 1.3 Main instruments

[0033] Centrifuge (Thermo), constant temperature metal bath (eppendorf), NanoDrop 2000 (Thermo), qPCR instrument (Roche), PCR instrument (ABI)

[0034] 1.4 Main method

[0035] 1.4.1 RNA extraction and real-time fluorescen...

Embodiment 2

[0041] Example 2: Effect of RNase R on the expression of hsa_circ_0003764.

[0042] 1. Materials, reagents, equipment

[0043] 1.1 Main reagents

[0044] RNase R (Epicentre), RNA Purification Kit (QIAGEN), Reverse Transcriptase (Takara), Qpcr Enzyme (Roche).

[0045] 1.2 Main instruments Constant temperature metal bath (eppendorf), qPCR instrument (Roche), PCR instrument (ABI), centrifuge (Thermo)

[0046] 1.3 Main method

[0047] 1.3.1 RNase R treatment of total RNA

[0048] Take 2 copies of 2.5 μg total RNA of the endometrial tissue of AS patients extracted by Trizol method, add 0.5 μl (10 U) of RNase R and enzyme-free water respectively, then add 2 μl 10x Reaction Buffer, and finally add enzyme-free water to form a 20 μl reaction system, metal Purification was performed after 15 minutes in the bath.

[0049] 1.3.2 RNA Purification Add enzyme-free water to 100 μl of the RNase R-treated sample, add 350ul RLT and mix well. Add 250 μl of absolute ethanol, pipette evenly, ...

Embodiment 3

[0054] Example 3: Localization of hsa_circ_0003764 in cells

[0055] 1. Materials, reagents, equipment

[0056] 1.1 Main reagents

[0057] Fluorescence in situ hybridization probe (Guangzhou Boxin Biotechnology Co., Ltd.), hybridization kit (Guangzhou Boxin Biotechnology Co., Ltd.), xylene, ethanol, DEPC water, PBS, proteinase K, formamide, SSC

[0058] 1.2 Main instruments

[0059] Hybridization instrument (Thermobrite), shaker, fluorescence microscope (Leica).

[0060] 1.3 Main method

[0061] The expression of hsa_circ_0003764 was detected by fluorescence in situ hybridization on paraffin sections. Xylene was treated 3 times, 5 min each time; washed 2 times with 100% alcohol, 5 min each time, and air-dried; prepare proteinase K mixture (take 20 μl Tris-Hcl, add 180 μl DEPC-treated water, and then add 0.2 μl proteinase K to make 200μl and mix well); add proteinase K mixture dropwise on the slice, cover with membrane and ensure even coverage, incubate at 55°C for 10-15mi...

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Abstract

The invention discloses an application of hsa_circ_0003764 as a marker to preparation of a diagnosis preparation for intrauterine adhesion. The expression of hsa_circ_0003764 in endometrium cicatricial tissue of patients suffering from intrauterine adhesion is notably increased, and the hsa_circ_0003764 can resist digestion of ribonucleic acid (RNA) excision enzymes R ( RNase R), and more stable than liner RNA. In vitro cell experiment proves that the hsa_circ_0003764 can cause endometrial stromal apoptosis and epithelial fibrosis. si-hsa_circ_0003764 has the effect of resisting epithelial fibrosis. Therefore, the hsa_circ_0003764 can be used as the marker for diagnosis of the endometrial stromal so as to prepare a diagnosis reagent for diseases caused by uterus fibrosis. The invention relates to an application of the reagent for detecting the hsa_circ_0003764 to preparation of a diagnosis reagent of the intrauterine adhesion.

Description

technical field [0001] The present invention relates to the use of hsa_circ_0003764 as a diagnostic marker for intrauterine adhesions and the preparation of drugs for targeted treatment of intrauterine adhesions. Background technique [0002] Intrauterine adhesions (IUA), or Asherman syndrome, is caused by a variety of factors that cause endometrial basal layer damage, functional layer regeneration and repair disorders, and the formation of intramural adhesions characterized by endometrial fibrosis. IUA is the most common cause of uterine infertility, accounting for as high as 25-30% of infertility patients; The incidence of fibrosis was significantly increased. At present, the main diagnosis is to observe the degree of endometrial fibrosis and adhesion under hysteroscopy, and the preferred treatment method is hysteroscopic adhesion dissection, and place intrauterine devices or balloons or biomaterials to block the anterior and posterior walls of the uterus to prevent Re-a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12N15/113A61K45/00A61P15/00
Inventor 胡娅莉赵光锋宋敏敏李若天
Owner NANJING DRUM TOWER HOSPITAL
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