| ObjectiveThe purpose is to explore and analyze the effect of secretions derived from mesenchymal stem cells overexpressing Fam20B(Family of Sequence Similarity 20 Member B)on the mechanism of bone defect healing.Methods⑴The results of the RT-qPCR experiment showed that after coculturing rat bone marrow mesenchymal stem cells with MSC-exo,Fam20 B MSC-exo,and Fam20 B control MSC-exo,the expression level of Fam20 B m RNA in the Fam20 B MSC-exo group cells was significantly higher than that in the MSC-exo group and Fam20 B control MSC-exo group(P<0.05),and the differences between the groups were statistically significant The ⑵RT-qPCR experiment results showed that compared with bone marrow mesenchymal stem cells transfected with MSC-exo and Fam20 B control MSC-exo,the expression levels of osteogenic related genes Runx2,Ocn,and BMP2 in bone marrow mesenchymal stem cells transfected with Fam20 B MSC-exo were significantly higher(P<0.05),and the differences between groups were statistically significant.⑶Western Blot experiments showed that compared with the Fam20 B MSC-exo group,the expression of osteogenic related proteins Runx2,Ocn,and BMP2 in bone marrow mesenchymal stem cells of rats in the MSC-exo group and Fam20 B control MSC-exo group was down regulated(P<0.05),with significant differences between the groups and statistical significance.⑷Micro CT results showed no significant changes in the blank control group;There was no significant difference in the repair of bone defects in the injury group rats,but the defect in the bone defect model was significant;The range of bone defects in the exosomes group decreased;The range of bone defects in the exosome+Fam20B group bone defect model was further reduced(P<0.05),and the differences between the groups were statistically significant.⑸The HE staining results showed that there was no significant change in the blank control group,while there was almost no new bone tissue formation in the bone tissue defect area of the injury group rats.Both the exosomes and exosomes+Fam20B groups had new bone tissue formation,but the new bone structure formed in the exosomes group was uneven,while the exosomes+Fam20B group formed more uniform new bone tissue,which was closer to normal bone tissue,and had more abundant bone mass.The results of Masson staining showed that there was no significant change in the blank control group,while the blue collagen component in the bone tissue of the injury group rat bone defect model was not very significant;The collagen fiber composition of bone tissue in the exosome-treated rat bone defect model increased,and some blue staining was weaker;In the exosome+Fam20B group of rat bone defect model,the collagen composition of bone tissue further increased,and the collagen blue staining was obvious.⑹ The RT-qPCR experiment results showed that compared with the injury group,the relative expression levels of osteogenic related genes Runx2,Ocn,and BMP2 in the new bone tissue of the exosome-treated rat bone defect model were significantly higher(P<0.05).Compared with the exosome-treated group,the relative expression levels of osteogenic related genes Runx2,Ocn,and BMP2 in the new bone tissue of the exosome-treated rat bone defect model were further increased(P<0.05),and the differences between the groups were statistically significant.⑺ Western Blot experiments showed that compared with the injury group,the expression levels of osteogenic related proteins Runx2,Ocn,and BMP2 in the new bone tissue of the exosome group rat bone defect model were significantly higher(P<0.05).The expression levels of osteogenic related genes Runx2,Ocn,and BMP2 in the new bone tissue of the exosome+Fam20B group rat bone defect model were significantly higher than those of the exosome group(P<0.05),and the differences between the groups were statistically significant.ResultsRT-qPCR experiments showed that after co-culture of BMSCs with MSC-exo,Fam20BMSC-exo,and Fam20 B control-MSC-exo,the expression levels of Fam20 B m RNA,Runx2,OCN,and BMP2 were higher in the cells of Fam20BMSC-exo group than in the control group(P < 0.05).Western Blot experiments showed that Fam20BMSC-exo group had the strongest expression of bone-related genes(P < 0.05).In the rat bone defect model,Micro-CT results showed that the extent of bone defects was significantly reduced in the exosome+Fam20B group compared with other groups(P < 0.05).HE staining showed that inflammatory cells were significantly reduced and new bone formation was increased in the exosome+Fam20B group.Masson staining results showed that the exosome+Fam20B group had well arrange and dense collagen fibers compared with the injury group.The collagen fibers were well arranged and dense compared to the damaged group.ConclusionFam20B loaded by exosomes can promote osteogenic gene expression and osteogenic differentiation of osteoblasts,and help repair bone defects. |
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