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A method to promote in vitro expansion of mouse retinal precursor cells by inhibiting microRNAs

A microRNA, mouse retina technology, applied in the field of stem cell application, can solve the problem of small number and achieve the effect of promoting expansion

Active Publication Date: 2021-02-09
FOURTH MILITARY MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the bottlenecks faced by the above-mentioned transplantation therapy are mainly biological safety and obtaining a sufficient number of RPCs
[0006] RPCs are derived from the embryonic retina, the number is small, and there are ethical restrictions on the sampling

Method used

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  • A method to promote in vitro expansion of mouse retinal precursor cells by inhibiting microRNAs
  • A method to promote in vitro expansion of mouse retinal precursor cells by inhibiting microRNAs
  • A method to promote in vitro expansion of mouse retinal precursor cells by inhibiting microRNAs

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Embodiment Construction

[0027] The present invention is described in further detail below in conjunction with accompanying drawing:

[0028] 1. Primary culture process: Prepare pregnant C57 mice at 17.5 days pregnant, cut out the fetal mice, move the fetal mice into a clean table, and place them in sterile PBS solution. Under the microscope, the fetal mouse eyeballs were peeled off, and the eyeballs were placed in the pre-prepared PBS droplet. The collected eyeballs were torn open with ophthalmic forceps, and the retinal tissue was taken out and put into fresh PBS droplet. Collect retinal tissue in a 1.5ml centrifuge tube. After centrifugation at 800rpm / min for 5min, the supernatant was discarded. Add trypsin digestion solution containing EDTA, digest at 37°C for 10 min, and add Neurobasal medium to terminate digestion. After centrifugation at 1300rpm / min for 5min, the supernatant was discarded. Cell pellets were resuspended in Neurobasal complete medium and plated on cell culture dishes. Add 2%...

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Abstract

The invention discloses a method for promoting the in vitro expansion of mouse retinal precursor cells by inhibiting microribonucleic acid. MicroRNA miR-449a-5p is a non-coding single-stranded RNA molecule of 22 nucleotides in length, and its coding DNA sequence is located on mouse chromosome 13. Retinal precursor cells are a class of cells with self-renewal ability and the potential to differentiate into retinal neurons and glial cells. By synthesizing the antisense strand of miR-449a-5p and using the principle of base pairing to interfere with the level of endogenous miR-449a-5p, it can effectively promote the expansion of retinal precursor cells in vitro. Since retinal precursor cells are the developmental source of photoreceptor cells in the mature neural retina, their efficient in vitro expansion method could facilitate cell therapy based on stem / precursor cell transplantation.

Description

Technical field: [0001] The invention belongs to the field of stem cell application, and in particular relates to a miR-449a-5p inhibitor sequence that can promote the expansion of mouse retinal precursor cells in vitro. Retinal precursor cells have important application potential in the damage repair of retinal diseases, so this expansion method can be applied to the treatment research of various retinal damage and degeneration models. In addition, because the sequence of mouse and human miR-449a is the same, this method is expected to be applied to the in vitro expansion of human retinal precursor cells. Background technique: [0002] microRNA is a kind of non-coding single-stranded RNA molecule encoded by endogenous genes with a length of about 22 nucleotides. Participate in post-transcriptional modification and regulate the expression of target mRNA. It can down-regulate the expression level of protein by promoting the degradation of mRNA or preventing the translation ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113C12N15/88C12N5/10
CPCC12N5/0621C12N15/113C12N15/88C12N2310/113C12N2510/00
Inventor 郑敏化郭辰峻韩骅
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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