| BACKGROUND: Bone regeneration has always been a hot issue in the field of orthopaedics,and the clinical need for effective bone regeneration therapy remains huge.With the deepening of tissue engineering research,the importance of mesenchymal stem cells in tissue regeneration has received more and more attention.The bone marrow-derived mesenchymal stem cells(BMSCs)are convenient for their acquisition methods.It is more commonly used.Some studies have pointed out that the supernatant of MSCs also has the function of repairing damaged tissues,and its effect may be related to the regulation mechanism of paracrine of stem cells.Exosome,one of the paracrine components of stem cells,plays a very important role in regulating the function of target cells by stem cells.Exosomes generally have a diameter of 40-160 nm and contain mRNA,microRNA,and protein components that are synthesized in the source cells.They enter the extracellular matrix through budding,and they are endocytosed by the target cells to release internal components and complete cell-to-cell communication.And exchange of information.Exosomes can effectively repair damaged tissues and organs such as heart,lung,kidney,and brain,indicating that transplanted exosomes can perform similar functions as transplanted stem cells.Therefore,we speculate that bone marrow mesenchymal stem cell-derived exosomes(BMSC-Exos)can exhibit similar biological functions as BMSCs,ie,promote bone regeneration.This topic will study the promoting effect and mechanism of bone marrow-derived mesenchymal stem cells exosomes on bone regeneration,hoping to provide theoretical basis for the future treatment of bone regeneration of exosomes,and also provide new clues for the clinical solution of bone regeneration.Ideas.OBJECTIVE: To investigate the characteristics of bone marrow-derived stem cell exosomes(BMSC-Exos)and to verify its ability to promote osteogenic differentiation through in vitro and in vivo experiments,as well as to explore its mechanism of promoting bone regeneration.Methods:(1)Isolation and culture of human bone marrow mesenchymal stem cells(BMSCs)to verify their surface antigen and multilineage differentiation potential;(2)Collection of BMSC-derived P4-P6 cell culture supernatants and extraction of BMSC-by kits Exos;(3)The morphological structure of BMSC-Exos was observed by transmission electron microscope.The surface antigen of BMSC-Exos was detected by immunoelectrophoresis.(4)Alizarin red and ALP staining confirmed the ability of BMSC-Exos to promote osteogenic differentiation in vitro;(5)The rat skull defect model was used to verify the ability of BMSC-Exos to promote bone regeneration in vivo;(6)The expression of genes and proteins related to osteogenic differentiation was detected by immunoelectrophoresis and qRT-PCR.RESULTS:(1)The morphology of human bone marrow mesenchymal stem cells(BMSCs)isolated and cultured was spindle-shaped,expressing CD29,CD44,and CD90,consistent with the characteristics of mesenchymal stem cells.(2)BMSC-Exos was a biconcave round or oval shape with a diameter of 40~120 nm(81.7±19.9).The surface antigen was consistent with BMSC.(3)Alizarin red staining and ALP staining results showed that the staining intensity of Exo+ osteogenic induction group was significantly higher than that of PBS+ osteogenic induction group.(4)X-ray and histological analysis of the rat skull defect model showed that the repair of the bone defect in the exosome group was better than the control group,and there were more new bone formation.(5)Immunoelectrophoresis showed that after treatment with exosomes,the content of OCN,Runx2,and β-catenin protein in MSC cells increased,and qRT-PCR showed that the gene expression of Runx2 and β-catenin was up-regulated.CONCLUSIONS: BMSC-Exos has the ability to promote bone regeneration and is associated with upregulation of the Wnt/β-catenin pathway. |
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