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LMO3 gene for regulating and controlling ADSCs osteogenic differentiation and tissue regeneration, and application thereof

A bone tissue regeneration and osteogenic differentiation technology, applied in the field of bone tissue regeneration, can solve problems that have not been reported

Pending Publication Date: 2021-03-09
辽宁省肿瘤医院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, there are no reports on the role of LMO3 in adipose-derived stem cells and its relationship with adipose-derived stem cell osteogenic differentiation and tissue regeneration.

Method used

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  • LMO3 gene for regulating and controlling ADSCs osteogenic differentiation and tissue regeneration, and application thereof
  • LMO3 gene for regulating and controlling ADSCs osteogenic differentiation and tissue regeneration, and application thereof
  • LMO3 gene for regulating and controlling ADSCs osteogenic differentiation and tissue regeneration, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Example 1 Isolation and cultivation of human adipose stem cells

[0067] Abdominal adipose tissue was collected during liposuction and stored in a 20mL sterile syringe for later use, and stored in ice packs. Adipose tissue was centrifuged in a 50mL centrifuge tube at 1000rpm / 5min to remove the lower liquid. Adipose tissue was digested with 0.2% type I collagenase at a ratio of 1:1 for 40 minutes on a shaker at 37°C. After standing still, it could be seen that it was divided into three layers. Remove the upper grease layer, fully shake and filter (200 mesh filter). Then centrifuge at 1000rmp / 5min for three times to remove the supernatant liquid, and the cell sediment therein can be seen. Pipette the cells evenly with cell culture medium. Cells were then seeded in culture dishes at a density of 5 × 10 5 / mL. The cells were then placed at 37°C, 5% CO 2 , 95% humidity incubator. Subculture to the third generation to obtain the third generation adipose stem cell hADSC...

Embodiment 2

[0068] Example 2 Joint analysis of GEO database and mRNA sequencing results

[0069] The series matrix files of GSE63754, GSE89330 and GSE72429 were downloaded from the GEO database (http: / / www.ncbi.nlm.nih.gov / geo / ). The platforms they are based on are GPL17077 (Agilent-039494SurePrint G3 Human GE v2 8x60K Microarray 039381), GPL16956 (Agilent-045997Arraystar Human lncRNA Microarray V3) and GPL16770 (Agilent-031181Unrestricted_Human_miRNA_V16.0_Microarray).

[0070] The GSE63754 dataset includes 3 ADSC osteogenic differentiation datasets and 3 control cell differentiation datasets. The GSE89330 dataset includes 4 datasets of osteogenic differentiation of ADSCs and 4 datasets of undifferentiated ADSCs. The GSE72429 dataset includes 4 datasets of ADSCs differentiated into osteoblast lineages and 4 datasets of undifferentiated cells from the same donor. mRNA sequencing was performed during hADSC osteogenic differentiation. By calculating the correlation between the two sample...

Embodiment 3

[0074] Example 3 Cell Functional Verification of LMO3 in ADSCs Osteogenic Process

[0075] 1. The adipose-derived stem cells were induced to osteoblast 0d, 7d, and 14 days respectively, RNA was extracted, and the expression trend of LMO3 at different times was detected by RT-qPCR. RT-q PCR was used to detect the expression changes of LMO3 after osteogenic induction of adipose-derived stem cells on day 0, day 7, and day 14 as follows: Image 6 As shown, the expression of LMO3 increases over time during osteogenic differentiation.

[0076] 2. Design and transfection of small interfering RNA sequences

[0077] Primers were designed for the LMO3 sequence. The primers were synthesized and purified by Shanghai Gemma Biotechnology Co., Ltd., and small interfering RNA sequences were designed to inhibit the expression of LMO3.

[0078] According to the starting and ending points of the human LMO3 sequence as its splicing point, design the best si RNA target point of the circ RNA base...

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Abstract

The invention relates to an LMO3 gene for regulating and controlling ADSCs osteogenic differentiation and tissue regeneration, and application thereof, in particular to an LMO3 gene, wherein the GeneID of the LMO3 gene is hsa-55885; and the nucleotide sequence of the LMO3 gene is shown as SEQ ID NO:1. The gene is specifically applied to the aspects of corresponding ADSCs osteogenic differentiation control, tissue regeneration products, regeneration regulation and the like. According to the invention, in-vitro and in-vivo experiments for regulating and controlling ADSCs osteogenic differentiation and tissue regeneration through the LMO3 prove that the LMO3 plays an important regulating and controlling role in the adipose derived stem cell osteogenic differentiation and / or tissue regeneration process; the LMO3 can be used as a potential target for tissue regeneration treatment; new LMO3 related drugs or factors are promoted to be developed and finally applied to the field of regenerative medicine; and real precise medical treatment in clinic is achieved.

Description

Technical field: [0001] The invention belongs to the technical field of bone tissue regeneration, and in particular relates to regulating ADSCs osteogenic differentiation and tissue regeneration LMO3 gene and application thereof. Background technique: [0002] Wound repair and reconstruction includes repair of trauma caused by skin or soft tissue injury, healing of chronic refractory wounds, repair of bone trauma, angiogenesis, repair of spinal cord, etc. Traditional autologous transplantation causes iatrogenic defects in donor tissue, which seriously affects the healing and quality of life of patients. The application of allogeneic transplantation is also limited by immune rejection and infection. Therefore, a new therapeutic strategy with limited secondary trauma, low immunogenicity, and ideal functional recovery is urgently needed. Adipose Derived Stem Cells (ADSCs) have attracted extensive attention in recent years. Adipose stem cells have a wide range of sources, mor...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/12C12Q1/6881A61K38/17A61K31/713A61L27/38A61P19/08
CPCC07K14/4702C12Q1/6881A61K31/713A61L27/3834A61P19/08C12Q2600/158A61K38/00
Inventor 康悦姜大庆
Owner 辽宁省肿瘤医院
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