Applicationof let-7c/let-7d in intrauterine adhesion and/or thin endometriumdiagnosis and treatment
A technology of let-7d and intrauterine adhesions, applied in the direction of medical preparations containing active ingredients, measurement/testing of microorganisms, biochemical equipment and methods, etc., to achieve the effect of increasing the number of glands and the thickness of the endometrium
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Embodiment 1
[0023] Example 1 Expression of let-7a, let-7b, let-7c, let-7d, let-7e, let-7f and let-7g in endometrial tissues of patients with normal and severe intrauterine adhesions
[0024] 1. Materials, reagents, equipment
[0025] 1.1 Source of human endometrial tissue
[0026] Collect 11 cases of normal endometrial tissue and 11 cases of endometrial tissue from patients with severe intrauterine adhesion syndrome. When obtaining endometrial tissue, the follicle size was detected by ultrasound to ensure that the endometrial tissue acquisition stage was unified into the late stage of proliferation. All participants signed a written informed consent form, which was approved by the Ethics Committee of Nanjing Drum Tower Hospital.
[0027] 1.2 Main reagents
[0028] Trizol (Invitrogen), reverse transcriptase (Takara), Qpcr enzyme (Roche), let-7 family member reverse transcription and qPCR primers.
[0029] 1.3 Main instruments
[0030] qPCR instrument (Roche), PCR instrument (ABI)
[0...
Embodiment 2
[0036] Example 2 Changes of fibrosis markers in endometrial epithelial cells after let-7c / let-7d reduction.
[0037] 1. Materials, reagents, equipment
[0038] 1.1 Main reagents
[0039] Collagenase (Sigma), Hyaluronidase (Sigma), DNAase (Roche), Epithelial Cell Culture Medium (Gibco), Serum (Gibco), Trizol (Invitrogen), Reverse Transcriptase (Takara), QPCR Enzyme (Roche), let-7c / 7dinhibitor (Ribobio, Guangzhou).
[0040] 1.2 Main instruments
[0041] Cell incubator, shaker, qPCR instrument (Roche), PCR instrument (ABI)
[0042] 1.3 Main method
[0043] 1.3.1 Isolation and culture of primary endometrial epithelial cells
[0044] Place the endometrial tissue in a new 60mm petri dish, cut the tissue until no lumps are visible to the naked eye, add the prepared digestive solution, blow and mix, and place it in a 37°C incubator for 5 minutes; take out the petri dish and observe the tissue under a microscope Digestion: Add DNase at a concentration of 4mg / mL, continue to diges...
Embodiment 3
[0049] Example 3 After let-7c / let-7d intervene in a mouse model of endometrial fibrosis, endometrial fibrosis changes.
[0050] 1. Materials, reagents, equipment
[0051] 1.1 Main reagents
[0052] HE staining solution (Solarbio), Masson staining solution (Thermo), angomir let-7c / let-7d and angomir control (Guangzhou Ruibo).
[0053] 1.2 Main instruments
[0054] Tissue embedding machine, slicer
[0055] 1.3 Main method
[0056] 1.3.1 Establishment of mouse endometrial fibrosis model
[0057] Select BALB / c female mice of 8-10 weeks old, weighing about 20 g, in estrus. The mice were anesthetized by intraperitoneal injection of 0.2ml of 4% chloral hydrate, and the limbs were fixed with medical adhesive tape. Cut the abdomen with surgical scissors, insert a self-made scraper into the upper part of the uterus, and scrape back and forth on one side of the uterus for about 2 minutes, resulting in rough endometrial surface; Intrauterine injection of 20 μl 2.5mg / ml LPS with insu...
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