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Application of gene SHISA4 as intervertebral disc degenerative change diagnosis and treatment marker

A technology for degenerative diseases and intervertebral discs, which can be applied in gene therapy, disease diagnosis, microbial measurement/testing, etc., and can solve the problems that the pathogenesis has not been clearly elucidated

Active Publication Date: 2016-03-16
BEIJING MEDINTELL BIOMED CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Surgical treatment for degenerative disc disease is risky, and early interventional treatment can achieve better curative effect. In addition, the specific pathogenesis of the disease has not yet been clearly elucidated

Method used

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  • Application of gene SHISA4 as intervertebral disc degenerative change diagnosis and treatment marker
  • Application of gene SHISA4 as intervertebral disc degenerative change diagnosis and treatment marker
  • Application of gene SHISA4 as intervertebral disc degenerative change diagnosis and treatment marker

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Example 1 Difference of SHISA4 gene expression in degenerative disc tissues and normal control tissues

[0062] 1. Sample collection

[0063] 40 cases of normal human intervertebral disc tissue were taken from patients with traumatic spinal burst fractures. Take 40 cases of intervertebral disc tissue from patients with degenerative disc disease, with an average age of 75 years. According to the Gr1es [5] scoring standard, they are all severely degenerated. The patients and their family members agreed to the sample collection.

[0064] 2. Extraction of nucleus pulposus tissue

[0065] Normal nucleus pulposus tissue: The normal intervertebral disc tissues were removed completely by the operation, and the white fibrous ring on the periphery and the jelly-like nucleus pulposus tissue in the center were visible. Soak the intervertebral disc with normal saline containing double antibody (penicillin-streptomycin) for 10 minutes. Use a curette to gently separate the nucleus pulposus ...

Embodiment 2

[0089] Example 2 Interference with SHISA4 gene expression

[0090] 1. siRNA design and synthesis

[0091] SiRNA sequence against SHISA4:

[0092] siRNA1-SHISA4:

[0093] The sense strand is 5'-UGUCUUUACAGCUUUAAUCUA-3' (SEQIDNO.3);

[0094] The antisense strand is 5'-GAUUAAAGCUGUAAAGACAUA-3' (SEQIDNO.4),

[0095] siRNA2-SHISA4:

[0096] The sense strand is 5'-UCUUUAGGAAGCCUUUAGGAA-3' (SEQIDNO.5);

[0097] The antisense strand is 5'-CCUAAAGGCUUCCUAAAGACA-3' (SEQIDNO.6),

[0098] siRNA3-SHISA4:

[0099] The sense chain is 5'-UGUUGUAGAGCUGUUGAUCCC-3' (SEQIDNO.7);

[0100] The antisense strand is 5’-GAUCAACAGCUCUACAACAGC-3’ (SEQIDNO.8)

[0101] Negative control siRNA sequence (siRNA-NC):

[0102] The sense strand is 5'-CGUACGCGGAAUACUUCGA-3' (SEQIDNO.9);

[0103] The antisense strand is 5'-UCGAAGUAUUCCGCGUACG-3' (SEQ ID NO. 10).

[0104] 2. Cell culture

[0105] Put the intervertebral disc nucleus pulposus tissue into a 100ml beaker containing 10m12% type II collagenase, and stir for about 60 minutes w...

Embodiment 3

[0122] Example 3 Effect of SHISA4 gene expression on the proliferation of nucleus pulposus cells

[0123] 1. Use CellCountingkit-8 (cck-8) kit to detect the proliferation of nucleus pulposus cells

[0124] 1.1 steps

[0125] The normal nucleus pulposus cells were cultured and transfected according to the method of Example 2. The cells were divided into three experimental groups:

[0126] Group 1: Cells were transfected with siRNA-NC as a control group;

[0127] Group 2: Cells were transfected with siRNA1-SHISA4.

[0128] 24h after transfection, 2×10 5 / ml density inoculated in 96-well cell culture plates, each experimental group designed three multiple wells, each well 100μl, placed in 37 ℃, 5% CO 2 After incubating in the incubator for 24 hours, add 10μl of CK-8 solution to each well of the culture wells to be tested, and incubate for 1 hour in the cell incubator, and measure the absorbance (OD) of each well at 450nm.

[0129] 1.2 Statistical methods

[0130] The experiment was repeated 3...

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Abstract

The invention discloses a gene SHISA4 and an expression product thereof. The gene SHISA4 can serve as a molecular marker for diagnosing and treating the intervertebral disc degenerative change. By detecting the gene SHISA4 and the expression product thereof in intervertebral disc tissue of a subject, whether the subject gets the intervertebral disc degenerative change or not is judged or whether the risk of getting the intervertebral disc degenerative change exists or not in the subject is diagnosed. By studying growth, senium and apoptosis indexes of nucleus pulposus cells cultured in vitro, it is proved that the gene SHISA4 serves as a drug target for treating the intervertebral disc degenerative change.

Description

Technical field [0001] The present invention relates to the field of biotechnology, in particular to the use of human SHISA4 gene in the diagnosis and treatment of degenerative disc disease. Background technique [0002] Degenerative discdisease (Degenerativediscdisease), manifested by symptoms such as reduced intervertebral disc space, herniation, compression, and sealing, is closely related to back pain, and the disease has caused a serious socioeconomic burden. Surgical treatment for degenerative disc disease has greater risks, and early interventional treatment can achieve better results. In addition, the specific pathogenesis of the disease has not been clearly elucidated. Therefore, looking for specific molecular markers related to the early diagnosis and prognosis of intervertebral disc degeneration has far-reaching significance for the realization of early diagnosis and individualized treatment of intervertebral disc degeneration. Summary of the invention [0003] In orde...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N33/68A61K45/00A61K48/00A61K38/17A61P19/08
CPCA61K38/17A61K45/00A61K48/00C12Q1/6883C12Q2600/158G01N33/6893G01N2800/10
Inventor 杨承刚孙耀兰
Owner BEIJING MEDINTELL BIOMED CO LTD
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