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Biomarker for diagnosing abortion and application thereof

A marker and biological technology, applied in the field of biomarkers for diagnosing abortion, can solve the problems of limited evaluation value and limited accuracy of clinical diagnosis and prognosis of threatened abortion, and achieve the effect of promoting self-renewal ability.

Pending Publication Date: 2022-03-11
DALIAN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Limited accuracy due to pulsatile progesterone secretion
There are also estrogen and inhibin A, etc. The above indicators are related to miscarriage, but they have limited value in the clinical diagnosis and prognosis of threatened abortion. Therefore, it is necessary to explore new indicators combined with the above indicators for abortion, which has a certain degree of research significance

Method used

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  • Biomarker for diagnosing abortion and application thereof
  • Biomarker for diagnosing abortion and application thereof
  • Biomarker for diagnosing abortion and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Clinical samples (villous tissue) from patients with inevitable abortion and healthy women with normal pregnancy, embedded in paraffin.

[0024] The expressions of Manα1, 3Man and ALG3 in villi tissue were detected.

[0025] 1. Immunohistochemical detection of the localization and expression of Manα1,3Man and ALG3 in villi tissue

[0026] (1) Dewaxing: put the prepared paraffin sections into the following solutions in turn: soak in xylene I for 10 minutes, soak in xylene II for 10 minutes, soak in 100% ethanol I for 5 minutes, soak in 100% ethanol II for 5 minutes, soak in 95% ethanol for 5 minutes , soak in 85% ethanol for 5 minutes, soak in 70% ethanol for 5 minutes; then rinse with slow water for 10 minutes. Dip in PBS 3 times, 5min each time.

[0027] (2) Antigen retrieval: Put the slices into citrate buffer and thaw in a microwave oven for 20 minutes. Take it out and let it cool down to room temperature naturally.

[0028] (3) Rinse with PBS 3 times, 5 min each...

Embodiment 2

[0047] Example 2: Detection of Manα1,3Man expression in trophoblast cells by Lectin staining and cell immunofluorescence

[0048] Slip-up: After the trophoblast cells were digested with trypsin, centrifuge at 800 rpm for 4 minutes, resuspend in fresh medium, and put the cell suspension into a petri dish with slides.

[0049] Collection: remove the culture medium in the culture dish, wash with PBS 3 times, 3 minutes each time.

[0050] Fixation: add 4% paraformaldehyde to the petri dish and fix it for 20min.

[0051] Paraformaldehyde was discarded, washed 3 times with PBS, 3 min each time.

[0052] Blocking: block with immunostaining blocking solution for 1h.

[0053] Primary antibody incubation: overnight at 4°C (rabbit anti-human ALG3 1:150; lectin GNA 1:300). Wash with PBS 6 times, 5min each time.

[0054] Fluorescently labeled secondary antibodies were added dropwise, incubated at room temperature for 1 h, and washed 6 times with PBS for 5 min each time.

[0055] DAPI ...

Embodiment 3

[0058] Example 3: Spheroid formation test to detect the self-renewal ability of trophoblast cells

[0059] (1) The trophoblast cells in good growth state were digested and centrifuged, the medium containing serum was removed, and washed twice with PBS.

[0060] (2) Prepare stem cell medium to resuspend cells (10 4 cells): DMEM / F12+1ⅹB27+20ng / mL bFGF+20ng / mL EGF.

[0061] (3) Cell plating: Choose a low-adhesion 6-well plate, with 4 ml of medium per well. The cultivation was completed in about 10 days, and the state of ball formation was observed.

[0062] Depend on image 3 It can be seen that transfection of ALG3 siRNA can inhibit the self-renewal ability of trophoblast cells and the stemness of trophoblast cells; while ALG3 cDNA can promote the self-renewal ability of trophoblast cells and the stemness of trophoblast cells.

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Abstract

The invention discloses a biomarker for diagnosing abortion and application thereof, and belongs to the technical field of biology. According to the application disclosed by the invention, high expression of alpha 1, 3 mannosylation (alpha 1, 3 mannosylation, Man alpha 1, 3 Man) and alpha 1, 3 mannosyl transferase (ALG3) in villus tissues of inevitable abortion patients is found for the first time, the Man alpha 1, 3 Man and alpha 1, 3 mannosyl transferase (ALG3) are taken as biological markers for diagnosing abortion, and meanwhile, it is proved that inhibition of expression of the Man alpha 1, 3 Man and alpha 1, 3 mannosyl transferase (ALG3) can promote differentiation capacity of embryonic trophoblast cells; placenta development is promoted, and the occurrence rate of abortion is reduced. Therefore, prediction and diagnosis of abortion are facilitated, and the placenta development mechanism is deeply understood. And a new theoretical basis is provided for clinical abortion diagnosis and treatment taking sugar as a target spot.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a biological marker for diagnosing abortion and its application. Background technique [0002] Sustaining a pregnancy requires proper development of the embryo and placenta. When the embryo implants, the trophoblast cells undergo self-proliferation while simultaneously differentiating into villous or extravillous trophoblast cells. In the human placenta, there are three major trophoblast subpopulations: cytotrophoblast (CTB), extravillous cytotrophoblast (EVT), and syncytiotrophoblast (STB). In the process of trophoblast differentiation, CTB cells have stem cell function and can undergo two different differentiations, one is to fuse into multinucleated STB cells, and the other is to undergo epithelial-mesenchymal transition to EVT cells. The STB, the outer layer of the placental villi, is the main site of gas and nutrient exchange between the mother and fetus. It secre...

Claims

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Application Information

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IPC IPC(8): G01N33/573G01N33/68G01N33/53
CPCG01N33/573G01N33/6893G01N33/5308G01N2800/36G01N2333/91102G01N2440/38G01N2400/00
Inventor 刘帅燕秋崔馨元王浩
Owner DALIAN MEDICAL UNIVERSITY
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