Method for differentiating synovium mesenchymal stem cells (SMSCs) to chondrocytes through in-vitro adenovirus mediated BMP-2/7 gene coexpression induction

A BMP-2, mesenchymal stem cell technology, applied in the field of biomedicine, can solve the problems of high price, easy degradation and inactivation

Inactive Publication Date: 2013-06-05
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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Problems solved by technology

[0004] The object of the present invention is to provide a method for inducing synovial mesenchymal stem cells to differentiate into chondrocytes through the co-expression of BMP-2 / 7 gene mediated by adenovirus in vitro. The adenovirus mediated BMP-2 / 7 The method of gene co-expression to induce the differentiation of synovial mesenchymal stem cells into chondrocytes needs to solve the technical problems of the prior art methods of inducing SMSCs to differentiate into chondrocytes, which are expensive, easy to degrade and inactivate

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  • Method for differentiating synovium mesenchymal stem cells (SMSCs) to chondrocytes through in-vitro adenovirus mediated BMP-2/7 gene coexpression induction
  • Method for differentiating synovium mesenchymal stem cells (SMSCs) to chondrocytes through in-vitro adenovirus mediated BMP-2/7 gene coexpression induction
  • Method for differentiating synovium mesenchymal stem cells (SMSCs) to chondrocytes through in-vitro adenovirus mediated BMP-2/7 gene coexpression induction

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Embodiment 1

[0030] Embodiment 1:1 materials and methods

[0031] 1.1 Experimental animals and main reagents and instruments

[0032] Six 3-month-old SPF grade New Zealand white rabbits, male or female, with an average weight of 2.1±0.3kg; 2-week-old SPF grade NOD / SCID nude mice, male or female, 20.4±0.5g; all provided by Slack, Chinese Academy of Sciences Provided by Experimental Animal Center.

[0033] H-DMEM, 2.5g / L trypsin / EDTA (GIBCO / BRL company, USA); dexamethasone, ascorbic acid-2-phosphate, L-proline, pyruvate (Sigma company, USA); TGF-β 3 , Insulin (insulin), ITS+Premix (R&D company, USA); FBS (HyClone company, USA); Trizol (Invitrogen company, USA); Oligo dt, dNTP, DEPC and various PCR primers (Injun Biotechnology Co., Ltd., China); reverse transcriptase MmLV (Fermentas, Canada); rTaq DNA synthetase, 100bp DNALadder (Takara, Japan); type I collagen, type II collagen IgG (Monosan, USA); CD34, CD44, CD45, CD90, CD14, CD19, CD73, CD105, CD29 and HLA-DR (human leukocyte DR antigen...

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Abstract

The invention relates to a method for differentiating SMSCs to chondrocytes through in-vitro adenovirus mediated BMP-2 / 7 gene coexpression induction. The method comprises the following steps: separating and culturing SMSCs; constructing a pAdTrack-CMV-BMP2-IRES-BMP7 double gene recombinant vector; transferring the fifth-generation SMSCs through utilizing the pAdTrack-CMV-BMP2-IRES-BMP7 double gene recombinant vector; amplifying the recombinant viruses obtained after the transfection; adding the recombinant viruses to a chondroblast induction culture solution for induction; and culturing for above 10 days to obtain fibrochondrocytes. The method which adopts the adenovirus mediated BMP-2 / 7 gene coexpression vector to transfer SMSCs has the advantages of high transgenic efficiency, instant expression and high safety, and allows the SMSCs to be differentiated to the chondrocytes through the in-vitro induction.

Description

Technical field: [0001] The present invention relates to the field of biomedicine, in particular to a method for differentiation of fibrocartilage-like cells, in particular to a method for inducing synovial mesenchymal stem cells to differentiate into chondrocytes through adenovirus-mediated BMP-2 / 7 gene co-expression in vitro. Background technique: [0002] MSCs (Mesenchymal Stem Cells) are derived from mesoderm tissue in the embryonic period. They are mesenchymal progenitor cell-like clone cells with high proliferative ability and multi-directional differentiation potential. Cells and other terminal cells have the ability of directed differentiation, showing broad application prospects in the fields of tissue engineering and regenerative medicine. With the continuous improvement of separation technology, people have been able to isolate and identify MSCs from various tissues including bone marrow, synovium, periosteum, skeletal muscle, and fat. In 2001, De Bari et al. suc...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10C12N15/861
Inventor 符培亮张雷吴海山吴宇黎丛锐军陈松
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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