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The role of the interaction between the hippo-yap pathway and the wnt pathway in the fate selection of adipose stem cells

A technology for the application of adipose stem cells, which is applied in the application field of the interaction between the Hippo-YAP pathway and the Wnt pathway in the selection of the fate of the adipose stem cell lineage, which can solve the problem of limited action time, failure to essentially promote the osteogenic differentiation of adipose stem cells, and reduce the The advantages of adipose-derived stem cells and the advantages of convenient use can achieve the effect of inhibiting adipogenic differentiation and promoting osteogenic or chondrogenic differentiation of adipose-derived stem cells

Active Publication Date: 2022-08-05
NANNING SECOND PEOPLES HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, there are mainly five methods for regulating the differentiation of adipose-derived stem cells: one is to select adipose-derived stem cells with strong osteogenic ability from the stem cell population by means of fluorescence-labeled flow cytometry and then use them for expansion. One-time use of adipose stem cells requires fluorescent labeling and flow cytometry screening, which is time-consuming and laborious, which reduces the advantages of adipose-derived stem cells and ease of use; the second is to use osteogenic induction fluid to induce osteogenesis of adipose-derived stem cells, and then apply them to tissues To promote the healing of bone defects on engineered bone, this method only exerts the osteogenic ability of adipose-derived stem cells in vitro in advance, and does not essentially promote the osteogenic differentiation of adipose-derived stem cells, and has no effect on the adipogenic differentiation of adipose-derived stem cells; The third is to use BMP and other cytokines that can promote the osteogenic differentiation of adipose-derived stem cells to directly add to the culture medium or play a role on adipose-derived stem cells in vivo in various slow-release ways, which can essentially improve the osteogenesis of adipose-derived stem cells differentiation ability, but its action time is limited, and it is difficult to maintain a stable and long-lasting effect of promoting osteogenesis or inhibiting adipogenesis; the fourth is to use small molecular compounds, such as dexamethasone and statins, to intervene in adipose-derived stem cells to improve their osteogenic ability. The first method is relatively simple and less costly, but the effect is limited; the fifth is to promote adipose osteogenic differentiation and inhibit adipogenic differentiation through genetic engineering, but the operation is cumbersome and expensive, and it is difficult to be widely promoted clinically.

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  • The role of the interaction between the hippo-yap pathway and the wnt pathway in the fate selection of adipose stem cells
  • The role of the interaction between the hippo-yap pathway and the wnt pathway in the fate selection of adipose stem cells
  • The role of the interaction between the hippo-yap pathway and the wnt pathway in the fate selection of adipose stem cells

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Experimental program
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Effect test

Embodiment 1

[0056] 1. Experimental materials

[0057] 1. Animals

[0058] 2-week-old New Zealand white rabbits (stem cells isolated from adipose tissue around the groin and epididymis) were obtained, provided by the Experimental Animal Center of Guangxi Medical University.

[0059] 2. Main reagents and instruments

[0060] Trizol kit was purchased from Bictrin Company in the United States, UV spectrophotometer, micropipette, sterilized ultra-thin PCR reaction tube were purchased from Sigma Company in the United States, mouse anti-human β3 integrin IgG was purchased from Chemicon Company, RPMI1640 (GIBICO Company), Fetal bovine serum (Gibco company), various digestive enzymes (packed by Sigma company), reverse transcription reagent (MBI company), PEGeneAmp PCR System 2400 (BD company, USA), SYBRGreen I quantitative PCR kit (Invetrgen company and Toyobo company) , US BD FACSCanto flow cytometer (US BD company), ABI-7300 quantitative PCR instrument. SPECT is Siemens DIACAM / VAX3300; FITC, ...

Embodiment 2

[0095] Example 2 Micromagnetic resonance elastography (μMRE) technique was used to observe the fracture healing at the fracture site.

[0096] a. Establishment of animal stress model: 2-week-old New Zealand white rabbits were taken. After pentobarbital anesthesia, a right anterolateral longitudinal incision was taken to separate the tibia, and the middle and lower tibia were treated to fracture, and then fixed with fine Kirschner wires. As a blank stress control; after anesthesia, the right anterolateral longitudinal incision was taken to separate the tibia under aseptic conditions, and the middle and lower tibia were treated to fracture, and about 0.3 cm of the tibia was taken out to create a tibial defect. After internal fixation with Kirschner wire, the fat around the groin and epididymis was taken. Bone regeneration in acanine cranial model using allogeneic adipose derived stem cells and coralscaffold. Biomaterials. 2013. 34(11) : 2655-64.), sutured after reduction, and es...

Embodiment 3

[0099] Example 3 Based on the results of μMRE imaging, the adipose stem cells at the fracture end were isolated and adipose stem cells were cultured in vitro.

[0100] a. Mixed cell acquisition: 2-week-old New Zealand white rabbits were obtained from adipose stem cells around the inguinal and epididymis;

[0101] b. Magnetic activated cell sorting (MACS): using the specific surface markers CD24a and CD200, the adipose stem cells in the mixed cell suspension were separated by the immunomagnetic bead sorting system (Miltenyi, Germany), cultured in vitro and Its surface markers were identified and confirmed by flow cytometry; (refer to the method of Belluoccio D et al.: J Bone Miner Res, 2010, 25(6): 1267-81)

[0102] Flow cytometry showed that rat ASCs positively expressed CD29 and CD44 molecules, and negatively expressed CD34, CD45 and CD31 molecules.

[0103] c. Detection of biological characteristics: observe cell proliferation, and use immunohistochemistry or fluorescence s...

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Abstract

The invention discloses the use of the interaction between the Hippo-YAP pathway and the classical Wnt in the preparation of a product for influencing the selection of adipose stem cell cell spectrum and fate, and specifically discloses the interaction between the YAP protein and β-catenin in the preparation of a product for affecting the adipose stem cell Use in products of cell profile fate selection. The present invention provides the use of a substance that inhibits the mutual binding of YAP protein and β-catenin in the preparation of a product for promoting the transfer of β-catenin to the nucleus, or the use of a substance that inhibits the mutual binding of YAP protein and β-catenin in the preparation of a product for improving the transcriptional activity of TCF Use in the product in , or use of a substance that inhibits the mutual binding of YAP protein and β-catenin in the preparation of a product for promoting the canonical Wnt pathway in the nucleus.

Description

technical field [0001] The invention belongs to the technical field of cell biology, in particular to the use of a Hippo-YAP pathway interacting with classical Wnt in the preparation of a product for influencing the selection of adipose stem cell cell spectrum fate, and more particularly to the mutual combination of YAP protein and β-catenin Use in the manufacture of a product for influencing adipose stem cell proliferation and cell lineage fate selection. Background technique [0002] With the rapid development of road traffic and mechanized production, fractures caused by high-energy trauma such as violence, such as comminuted fractures and postoperative nonunion, are the most difficult problems in orthopaedics. In clinic, autologous bone or allogeneic bone is usually used. transplantation to treat such diseases. However, autologous bone transplantation damages the site of bone extraction and the amount of bone obtained is limited. Allogeneic bone transplantation brings r...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/077
CPCC12N5/0653C12N5/0654C12N5/0655C12N2506/1384C12N2501/999
Inventor 李兴艳李军最黄克李林张波胡敏杨业静黄家志蔡敏杜勇军郑泽文周松马鸳霞李加洪宋菲
Owner NANNING SECOND PEOPLES HOSPITAL