Application of m6a modification-related gene alkbh5 in promoting nerve axon repair

A technology of injury repair and axon, applied in the field of biomedical research, can solve the problems of loss of nerve function, incomplete elucidation, and difficulty in the treatment of long-distance peripheral nerve injury, and achieve the effect of promoting recovery and promoting growth

Active Publication Date: 2022-04-15
NANTONG UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, based on microsurgery, local peripheral nerve transplantation or the application of tissue engineering materials, the treatment of peripheral nerve injury has great limitations, mainly in the difficulty of long-distance peripheral nerve injury treatment and partial loss of nerve function. The reason is that the key regulatory molecular targets and molecular mechanisms of regulation in nerve injury repair have not been fully elucidated

Method used

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  • Application of m6a modification-related gene alkbh5 in promoting nerve axon repair
  • Application of m6a modification-related gene alkbh5 in promoting nerve axon repair
  • Application of m6a modification-related gene alkbh5 in promoting nerve axon repair

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1. Detection of expression and localization of ALKBH5

[0044] (1), DRG sample protein extraction and Western blot

[0045] After the rat sciatic nerve crush model was prepared, the L4-L6 dorsal root ganglion (DRG) tissue on the side of the sciatic nerve crush was taken at 0h, 1d and 3d after operation, and the tissue protein sample was prepared by the tissue sample protein extraction kit: different time The spotted L4 and L5 DRG tissues were transferred from the cryovials to 1.5 ml RNase-free EP tubes with RNase-free tips. Add 150ul tissue lysate to the tube, add protease inhibitors, use a homogenizer to beat the tissue into a single-cell suspension, place it on ice for 15 minutes, and then centrifuge at 12000 rpm for 10 minutes at 4°C (involving protein sample preparation operation centrifugation temperature Both are 4). Carefully take out the centrifuge tube, draw the supernatant into a new EP tube, use the protein concentration detection kit to detect th...

Embodiment 2

[0049] Embodiment 2, the effect of interfering with the expression of ALKBH5 on the growth of neuron axons

[0050] (3) Isolation and culture of DRG neurons

[0051] Prepare the dissection solution and put it into a small dish, add double antibodies (penicillin and streptomycin) and place it on ice to pre-cool. Anesthetize 3 rats intraperitoneally, use surgical scissors to cut the skin from the tail along the spine to the head, carefully remove the entire spine, open the lamina from the neck, and use micro forceps to pull out all DRG tissues and put them into the dissection solution . After all the DRG tissues were taken out, the dissection fluid was discarded, and the tissues were rinsed twice with cell-grade PBS to remove excess tissues and blood. Discard the PBS, add 2ml of collagenase (3mg / ml), and transfer the tissue and digestion solution to a 5ml centrifuge tube. The tissue was fully shredded with micro scissors, placed in a cell culture incubator, and digested for...

Embodiment 3

[0056] Example 3, the effect of interfering with the expression of ALKBH5 on the growth of sciatic nerve axons

[0057] (6) Interfering with intrathecal injection of virus

[0058] Experimental animals SD rats are provided by the Experimental Animal Center, 15 per batch, adult male rats, about 200g. The experimental mice were randomly divided into 3 groups, 5 in each group. Before the operation, the rats were first anesthetized in the abdominal cavity, and then the rat’s back ilium was sterilized, and then the skin was exposed with surgical scissors, the muscle tissue was cut along the ilium with ophthalmic scissors, and the protruding spinous process was cut off with bone replacement scissors , and then use a small cotton ball dipped in normal saline to gently wipe off the blood and minced meat to clearly expose the intervertebral foramen of L4 and L5 DRG. The 2 / 8 type AAV virus packaged with shRNA was uniformly injected with a micro-injector. Injected into the interverte...

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Abstract

The invention discloses an application of m6A modification-related gene ALKBH5 in the preparation of medicines for treating diseases related to nerve axon damage. The invention also discloses an experimental verification of the application of m6A modification-related gene ALKBH5 in promoting the repair of nerve axon damage, which is characterized in that it comprises the following steps: S1, detecting the expression and location of ALKBH5; S2, interfering with the expression of ALKBH5 on nerve The effect of the axon growth; S3, the effect of interfering with the expression of ALKBH5 on the axon growth of the sciatic nerve; S4, the process of screening and functional testing of the ALKBH5 target gene LPIN2. The present invention verifies that interfering with ALKBH5 gene both in vivo and in vitro can significantly promote the growth of neuron axon, and promote the recovery of sciatic nerve function; and through transcriptome analysis and target gene verification after ALKBH5 interference, the target gene LPIN2 regulated by ALKBH5 is found and verified in Role in regulation of neuronal axon outgrowth.

Description

technical field [0001] The invention belongs to the technical field of biomedical research, and specifically relates to the application of m6A modification-related gene ALKBH5 in promoting nerve axon damage repair. Background technique [0002] With the development of human society, peripheral nerve injuries caused by wars, car accidents, and sports injuries have a relatively high incidence rate, which has brought great harm to patients and caused serious social and family burdens. At present, based on microsurgery, local peripheral nerve transplantation or the application of tissue engineering materials, the treatment of peripheral nerve injury has great limitations, mainly in the difficulty of long-distance peripheral nerve injury treatment and partial loss of nerve function. The reason is that the key regulatory molecular targets and molecular mechanisms of regulation in nerve injury repair have not yet been fully elucidated. Contents of the invention [0003] The tech...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K45/00A61P25/00G01N33/68
CPCA61K45/00A61P25/00G01N33/68
Inventor 于彬王东刘明稳顾晓松
Owner NANTONG UNIVERSITY
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