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Targets for diagnosis and treatment of degenerative disc disease

A technology for degenerative diseases and intervertebral discs, applied in bone diseases, microbiological determination/inspection, medical preparations containing active ingredients, etc., can solve the problems that the pathogenesis has not been clearly elucidated

Active Publication Date: 2019-01-29
QINGDAO MEDINTELL BIOMEDICAL 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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  • Targets for diagnosis and treatment of degenerative disc disease
  • Targets for diagnosis and treatment of degenerative disc disease
  • Targets for diagnosis and treatment of degenerative disc disease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 2

[0089] Embodiment 2 interferes with the expression of TULP2 gene

[0090] 1. siRNA design and synthesis

[0091] siRNA sequences against TULP2:

[0092] siRNA1-TULP2:

[0093] The sense strand is 5'-UCAUCAAUGUGUCAUUAUCCU-3' (SEQ ID NO.3);

[0094] The antisense strand is 5'-GAUAAUGACACAUUGAUGAGA-3' (SEQ ID NO.4),

[0095] siRNA2-TULP2:

[0096] The sense strand is 5'-AGGUAAUUAGAAGUUUUGCUC-3' (SEQ ID NO.5);

[0097] The antisense strand is 5'-GCAAAACUUCUAAUUACCUCA-3' (SEQ ID NO.6),

[0098] siRNA3-TULP2:

[0099] The sense strand is 5'-UCUUUUCCAAGUCUUCUUCCU-3' (SEQ ID NO.7);

[0100] The antisense strand is 5'-GAAGAAGACUUGGAAAAGAAG-3' (SEQ ID NO.8)

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

[0102] The sense strand is 5'-CGUACGCGGAAUACUUCGA-3' (SEQ ID NO.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 filled with 10ml2% ...

Embodiment 3

[0122] Example 3 Effect of TULP2 gene expression on proliferation of nucleus pulposus cells

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

[0124] 1.1 steps

[0125] The culture and transfection of normal nucleus pulposus cells were carried out according to the method of the previous embodiment 2, and the cells were divided into three experimental groups:

[0126] Group 1: Cells transfected with siRNA-NC, used as the control group;

[0127] Group 2: cells transfected with siRNA2-TULP2.

[0128] 24h after transfection, with 2×10 5 Inoculate in a 96-well cell culture plate at a density of / ml, design triplicate wells for each experimental group, 100 μl per well, place at 37°C, 5% CO 2 Incubate in an incubator. After 24 hours of cell attachment, add 10 μl of CK-8 solution to each well of the culture wells to be detected, continue to incubate in the cell incubator for 1 hour, and measure the absorbance value (OD value...

Embodiment 4

[0142] Example 4 Effect of TULP2 gene on apoptosis of nucleus pulposus cells

[0143]Use Annexin V-FITC / PI double staining method to detect cell apoptosis by flow cytometry, the steps are as follows:

[0144] 1. Cell transfection: The nucleus pulposus cells were transfected with siRNA2-TULP2 and siRNA-NC according to the method in Example 2.

[0145] 2. After 48 hours of cell transfection, the cells were harvested, and the cells were processed according to the operating instructions of the Annexin V-FITC / PI Double Staining Kit (purchased from Invitrogen).

[0146] 3. The cells treated in step 2 are detected by flow cytometry.

[0147] 4. Statistical methods

[0148] Statistical cell apoptosis rate, the experiment is repeated 3 times, the result data are expressed in the form of mean ± standard deviation, using SPSS13.0 statistical software for statistical analysis, the difference between the two Using t test, it is considered statistically significant when P<0.05.

[0149]...

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Abstract

The invention discloses a diagnosis and treatment target TULP2 gene of a degenerative disc disease. Whether a subject has the degenerative disc disease or not can be judged or whether the subject has the risk of the degenerative disc disease or not can be diagnosed by detecting the content of TULP 2 gene and an expression product thereof in intervertebral disc tissues of the subject. In addition, the TULP2 gene is proved to be used as a drug target for treating the degenerative disc disease by researching growth, senility and apoptosis indexes of the in-vitro cultured nucleus pulposus cells.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the application of human TULP2 gene in the diagnosis and treatment of intervertebral disc degeneration. Background technique [0002] Degenerative disc disease (Degenerative disc disease) is characterized by symptoms such as disc space shrinkage, herniation, extrusion, and sealing. It is closely related to back pain, and the disease has caused a serious social and economic burden. Surgical treatment for intervertebral disc degenerative lesions 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. Therefore, searching for specific molecular markers related to the early diagnosis and prognosis of intervertebral disc degenerative diseases has far-reaching significance for the early diagnosis and individualized treatment of intervertebral disc degenerative diseases. Con...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6883A61K45/00A61P19/08
CPCA61K45/00C12Q1/6883C12Q2600/158C12Q2600/178
Inventor 杨承刚李曙光孙耀兰
Owner QINGDAO MEDINTELL BIOMEDICAL CO LTD
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