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Method for promoting in-vitro amplification of RPCs (retinal progenitor cells) of mouse by inhibiting microRNA (micro ribonucleic acid)

A mouse retina and precursor cell 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: 2018-05-29
FOURTH MILITARY MEDICAL UNIVERSITY
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  • 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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  • Method for promoting in-vitro amplification of RPCs (retinal progenitor cells) of mouse by inhibiting microRNA (micro ribonucleic acid)
  • Method for promoting in-vitro amplification of RPCs (retinal progenitor cells) of mouse by inhibiting microRNA (micro ribonucleic acid)
  • Method for promoting in-vitro amplification of RPCs (retinal progenitor cells) of mouse by inhibiting microRNA (micro ribonucleic acid)

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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 in-vitro amplification of RPCs (retinal progenitor cells) of a mouse by inhibiting microRNA (micro ribonucleic acid). MicroRNA miR-449a-5p is a non-codedsingle-stranded RNA molecule with length of 22 nucleotides, and the coded DNA sequence of the molecule is located on chromosome 13 of the mouse. The RPCs are cells with self-renewal capacity as wellas potential of being differentiated into retinal neurons and neuroglia. By means of synthesis of an antisense strand of miR-449a-5p, interference to level of endogenous miR-449a-5p is realized according to the principle of complementary pairing of basic groups, and in-vitro amplification of the RPCs can be effectively performed. The RPCs are development sources of photoreceptor cells in mature neural retina, so that the effective in-vitro amplification method can promote 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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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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