Method for delaying senescence of mesenchymal stem cells through FOXP1 gene editing and mutation

Abstract

The invention relates to the technical field of cell biology, in particular to a method for delaying senescence of mesenchymal stem cells through FOXP1 gene editing and mutation. The method includes the following steps: (1) separating and culturing human umbilical cord mesenchymal stem cells; (2) adding plasmids, donor DNA (deoxyribonucleic acid) and lipidosome into the culture solution obtained in the step (1) for transfection; (3) transferring the human umbilical cord mesenchymal stem cells transfected in the step (2) into a human umbilical cord mesenchymal stem cell culture medium containing penicillin and streptomycin, and performing culturing; (4) carrying out screening culture on the human umbilical cord mesenchymal stem cells cultured in the step (3); and (5) carrying out digestion passage on the human umbilical cord mesenchymal stem cells cultured in the step (4), and performing amplifying until stably transferred cell strains are obtained. Compared with the prior art, the method has the advantages that the proliferation speed of the mesenchymal stem cells can be obviously promoted by one time, and the cell senescence process can be delayed by 50%.

Description

technical field [0001] The invention relates to the technical field of cell biology, in particular to a method for delaying the aging of mesenchymal stem cells through FOXP1 gene editing and mutation. Background technique [0002] Mesenchymal stem cells (MSCs) are non-hematopoietic adult pluripotent stem cells. Friedenstein first proposed the concept of MSCs to define a group of stem cells that can form fibroblast clonal colonies (CFU-F) in vitro , but also osteoblast, chondrocyte and adipocyte differentiation (Friedenstein et al., 1966). It has a high capacity for self-renewal while possessing the ability to differentiate into osteoblasts, chondrocytes, and adipocytes (Caplan, 2017). Most studies regard osteogenic, adipogenic and chondrogenic differentiation ability as the "gold standard" for defining mesenchymal stem cells (Bianco et al., 2008b). MSC cell surface markers include CD73, CD90, and CD105, but do not express CD11b, CD14, CD34, CD45, and HLA-DR. [0003] At p...

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Problems solved by technology

[0015] However, the above methods have the following problems: 1) use viral vectors, such as lentiviruses, or adenoviral vectors to transfect cells, provide donor DNA for CRISPR / Cas9, and carry out DNA recombination

The viral vectors used in these manipulations all carry the risk of random insertion into the cell genome or pose a risk to the operator during the experiment

2) Traditional gene editing and mutation, mediated by wild-type Cas9 gene, are prone to off-target effects and gene mutations, which affect cell safety

3) Using microRNA for cell transfection and treatment, 1-2 miRNAs alone are not effective, and combined use can easily lead to off-target effects

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