Construction Of Fibrin-targeting-exosome And Its Application In Bone Defect Repair

Bone repair is a very delicate and complicated process,involving the mutual interaction of cellular behaviors of niche cells in the bone environment.The complex signal molecules in the microenvironment regulate inflammation,revascularization,as well as mineralization and remodeling of bone tissue together.The study of mesenchymal stem cells(MSCs)has always been a research focus in the field of tissue repair.In recent years,researchers have found that stem cells exert their functions mainly through paracrine mechanism.Exosomes are important paracrine products,containing a variety of bioactive substances,and play an important role in the process of information transmission between cells and the regulation of cell behavior.With the deepening of research,exosomes have also faced problems such as low yield,easy removal by the system,and lack of cell targeting specificity,which may limit the therapeutic application of exosomes.Therefore,the functionalization of exosomes has become a research trend.This thesis attempted to use the hydrophobic interaction between the lipid chain of DMPE-PEG-CREKA and exosomal membrane lipid molecules to immobilize the CREKA fibrin targeting peptide on the surface of exosomes secreted from adipose-derived mesenchymal stem cells(Exos).Hopefully,the obtained CREKA functionalized exosomes(CREKA-Exos),can accumulate and reside in bone defects for a longer time and enhance the bone repair.This thesis studied the regulatory effects of Exos and CREKA-Exos on three key bone repair cells,including endothelial cells,macrophages,and bone marrow mesenchymal stem cells.The results showed that Exos and CREKA-Exos promoted endothelial cells to express vascular-related genes(eg.v WF,VEGF),induced their tube formation and promoted cell migration.They inhibited the expression of pro-inflammatory factors in pro-inflammatory(M1 type)macrophages,and stimulated the expression of anti-inflammatory factors,enhanced the phenotypic shift of macrophages from M1 type to M2 type.They also promoted the migration of bone marrow mesenchymal stem cells and induced their osteogenic differentiation by levelling up the expression of osteogenic genes(eg.ALP,RUNX2).In addition,this thesis used rat femoral defect as a model to studied the effect of CREKA-Exos on bone tissue repair in vivo.Results showed that the exosomes modified by CREKA could stay in the bone defect for more than two weeks and improved the bone repair process.In summary,this thesis used DMPE-PEG-CREKA to modify the surface of exosomes so that exosomes can target fibrin,thereby increasing the enrichment and retention of exosomes in tissue defects and promoting bone tissue repair.This thesis provided a new strategy to improve the efficiency of exosomes and showed that CREKA-Exos has great application value in the field of bone tissue repair.